The tumor microenvironment acidification confers treatment resistance; therefore, the interference with pH regulating systems is considered a new therapeutic strategy. In this study, two human prostate cancer cell lines, PC3 and LNCaP, have been treated in vitro with proton pump inhibitors (PPIs), namely Lansoprazole, Esomeprazole (V-ATPases-inhibitors), Cariporide, and Amiloride (NHE1-inhibitors). The cell viability and pH were assessed at several drug concentrations either at normoxic or hypoxic conditions. Since Esomeprazole showed the highest toxicity towards the PC3 cancer cells compared to LNCaP ones, athymic nude mice bearing subcutaneous or orthotopic PC3 tumors were treated with Esomeprazole (dose: 2.5 mg/kg body weight) for a period of three weeks—and tumor growth was monitored. MRI-CEST tumor pH imaging with Iopamidol was performed upon treatment at 3 h, 1 week (in combination with FDG-PET), and after 2 weeks for evaluating acute, early, and late responses. Although acute tumor pH changes were observed in vivo, long-term studies on both PC3 prostate cancer models did not provide any significant change in tumor acidosis or tumor growth. In conclusion, this work shows that MRI-CEST tumor pH imaging is a valuable tool for assessing the in vivo treatment response to PPIs.

Novel anticancer treatments target the pH regulating system that plays a major role in tumor progression by creating an acidic microenvironment, although few studies have addressed their effect on tumor acidosis. In this study, we investigated in vivo several proton pump inhibitors (PPIs) targeting NHE-1 (Amiloride and Cariporide) and V-ATPase (Esomeprazole and Lansoprazole) proton transporters in the DU145 androgen-insensitive human prostate cancer model. In cellulo results showed that DU145 are sensitive, with decreasing efficacy, to Amiloride, Esomeprazole and Lansoprazole, with marked cell toxicity both in normoxia and in hypoxia, with almost any change in pH. In vivo studies were performed upon administration of Esomeprazole to assess both the acute and chronic effects, and Iopamidol-based tumor pH imaging was performed to evaluate tumor acidosis. Although statistically significant tumor pH changes were observed a few hours after Esomeprazole administration in both the acute study and up to one week of treatment in the chronic study, longer treatment resulted in a lack of changes in tumor acidosis, which was associated to similar tumor growth curves between treated and control groups in both the subcutaneous and orthotopic models. Overall, this study highlights MRI-CEST tumor pH imaging as a valid approach to monitoring treatment response to PPIs.

Commercially available near-infrared (NIR) dyes, including indocyanine green (ICG), display an end-tail of the fluorescence emission spectrum detectable in the short-wave infrared (SWIR) window. Imaging methods based on the second NIR spectral region (1,000–1,700 nm) are gaining interest within the biomedical imaging community due to minimal autofluorescence and scattering, allowing higher spatial resolution and depth sensitivity. Using a SWIR fluorescence imaging device, the properties of ICG vs. heptamethine cyanine dyes with emission >800 nm were evaluated using tissue-simulating phantoms and animal experiments. In this study, we tested the hypothesis that an increased rigidity of the heptamethine chain may increase the SWIR imaging performance due to the bathochromic shift of the emission spectrum. Fluorescence SWIR imaging of capillary plastic tubes filled with dyes was followed by experiments on healthy animals in which a time series of fluorescence hindlimb images were analyzed. Our findings suggest that higher spatial resolution can be achieved even at greater depths (>5 mm) or longer wavelengths (>1,100 nm), in both tissue phantoms and animals, opening the possibility to translate the SWIR prototype toward clinical application.

Pancreatic ductal adenocarcinoma (PDAC) is highly metastatic and largely refractory to current therapies, underscoring the need to uncover the molecular drivers of progression to identify targetable vulnerabilities. In this study, we found that fibronectin type III domain–containing 4 (FNDC4), known for its role in macrophage polarization and metabolic regulation, was elevated in metastatic PDAC cells and correlated with poor patient outcomes. FNDC4 knockdown reduced tumor growth and metastasis in a diverse set of aggressive PDAC models. Mechanistically, FNDC4 enhanced cell cycle and apoptosis regulator 1 (CCAR1) stability, thereby sustaining CCAR1/β-catenin signaling. FNDC4 deficiency led to reduced CCAR1 and β-catenin expression and consequently impaired invasion and colony formation. Moreover, FNDC4 promoted immune evasion by driving macrophage polarization toward a protumorigenic M2 phenotype. FNDC4 loss shifted macrophage polarization toward an antitumor profile and increased CD4+ and CD8+ T-cell infiltration. Together, the effects of FNDC4 targeting resulted in reduced tumor burden, suppression of metastasis, and improved survival in immunocompetent murine PDAC models. Unexpectedly, FNDC4 localized to the nucleus, pointing to potential intranuclear activity. Transcriptomic and functional analyses further identified CCL5 as a critical downstream effector, required for recruiting CCR5+ T cells and mediating the immune effects of FNDC4 inhibition. Upstream, BHLHE40 directly activated FNDC4 transcription, which was stimulated by induction of epithelial–mesenchymal transition. Importantly, combining FNDC4 inhibition with claudin 18.2 chimeric antigen receptor T cells or chemotherapy resulted in enhanced tumor control compared with monotherapy. Together, these findings underscore the role of FNDC4 in promoting PDAC progression and the potential of FNDC4 as a target for innovative multimodal treatment strategies. Significance: FNDC4 is a key driver of pancreatic cancer invasiveness and immunosuppression that can be targeted to reprogram the pancreatic tumor microenvironment and suppress tumor metastasis, offering a promising therapeutic strategy.

Photodynamic therapy (PDT) is a technique based on the use of photosensitizers activated by light to destroy cancer cells in the presence of oxygen. This enables localized cancer treatment and, in some settings, fluorescence-guided visualization. However, the efficacy and clinical translation of PDT have been limited by the low specificity of traditional photosensitizers. The aim of the study is to create a ligand-guided PDT approach for pancreatic ductal adenocarcinoma (PDAC) using a peptide-conjugated photosensitizer binding to integrin αvβ6, which is a receptor linked to tumor growth and prevalent in PDAC cells. Current treatment options for this tumor are limited, with surgical resection and chemotherapy only effective when the tumor is detected early. Given the limited treatment options for PDAC, PDT via αvβ6 offers a new pathway for precision treatment. The cyclic peptide cyclo[FRGDLAFp(NMe)K], recognized for its high affinity to αvβ6, was chosen to guide a phthalocyanine-class photosensitizer toward αvβ6-expressing PDAC models. The PDT approach was further refined by developing 3D spheroid models and in vivo BxPc3 xenograft models in NOD/SCID mice, where its therapeutic efficacy was assessed. In the absence of a non-targeted control photosensitizer, a contribution from non-specific accumulation and EPR effects in the in vivo setting cannot be fully ruled out. This study highlights the potential of a peptide-guided photosensitizer, demonstrating uptake and photodynamic activity in spheroids, with moderate in vivo results addressing tumor microenvironment challenges. Optimization of PDT dosing, laser precision, and preclinical models, such as patient-derived xenografts, are crucial to enhance clinical translation.