Neoadjuvant chemotherapy can improve the survival of individuals with borderline and unresectable pancreatic ductal adenocarcinoma; however, heterogeneous responses to chemotherapy remain a significant clinical challenge. Here, we performed RNA sequencing (n = 97) and multiplexed immunofluorescence (n = 122) on chemo-naive and postchemotherapy (post-CTX) resected patient samples (chemoradiotherapy excluded) to define the impact of neoadjuvant chemotherapy. Transcriptome analysis combined with high-resolution mapping of whole-tissue sections identified GATA6 (classical), KRT17 (basal-like) and cytochrome P450 3A (CYP3A) coexpressing cells that were preferentially enriched in post-CTX resected samples. The persistence of GATA6^hi and KRT17^hi cells post-CTX was significantly associated with poor survival after mFOLFIRINOX (mFFX), but not gemcitabine (GEM), treatment. Analysis of organoid models derived from chemo-naive and post-CTX samples demonstrated that CYP3A expression is a predictor of chemotherapy response and that CYP3A-expressing drug detoxification pathways can metabolize the prodrug irinotecan, a constituent of mFFX. These findings identify CYP3A-expressing drug-tolerant cell phenotypes in residual disease that may ultimately inform adjuvant treatment selection.

Pediatric low-grade gliomas (pLGG) are the most common pediatric central nervous system tumors, with driving alterations typically occurring in the MAPK pathway. The ERK1/2 inhibitor ulixertinib (BVD-523) has shown promising responses in adult patients with mitogen-activated protein kinase (MAPK)-driven solid tumors. We investigated the antitumoral activity of ulixertinib monotherapy as well as in combination with MEK inhibitors (MEKi), BH3-mimetics, or chemotherapy in pLGG. Patient-derived pLGG models reflecting the two most common alterations in the disease, KIAA1549:BRAF-fusion and BRAF^V600E mutation (DKFZ-BT66 and BT40, respectively) were used for in vitro and in vivo (zebrafish embryos and mice) efficacy testing. Ulixertinib inhibited MAPK pathway activity in both models, and reduced cell viability in BT40 with clinically achievable concentrations in the low nanomolar range. Combination treatment of ulixertinib with MEKi or BH3-mimetics showed strong evidence of antiproliferative synergy in vitro. Ulixertinib showed on-target activity in all tested combinations. In vivo, sufficient penetrance of the drug into brain tumor tissue in concentrations above the in vitro IC₅₀ and reduction of MAPK pathway activity was achieved. In a preclinical mouse trial, ulixertinib mono- and combined therapies slowed tumor growth and increased survival. These data indicate a high clinical potential of ulixertinib for the treatment of pLGG and strongly support its first clinical evaluation in pLGG as single agent and in combination therapy in a currently planned international phase I/II umbrella trial.

Owing to their large size proteinaceous drugs offer higher operative information content compared to the small molecules that correspond to the traditional understanding of druglikeness. As a consequence these drugs allow developing patient-specific therapies that provide the means to go beyond the possibilities of current drug therapy. However, the efficacy of these strategies, in particular “personalized medicine”, depends on precise information about individual target expression rates. Molecular imaging combines non-invasive imaging methods with tools of molecular and cellular biology and thus bridges current knowledge to the clinical use. Moreover, nuclear medicine techniques provide therapeutic applications with tracers that behave like the diagnostic tracer. The advantages of radioiodination, still the most versatile radiolabeling strategy, and other labeled compounds comprising covalently attached radioisotopes are compared to the use of chelator-protein conjugates that are complexed with metallic radioisotopes. With the techniques using radioactive isotopes as a reporting unit or even the therapeutic principle, care has to be taken to avoid cleavage of the radionuclide from the protein it is linked to. The tracers used in molecular imaging require labeling techniques that provide site specific conjugation and metabolic stability. Appropriate choice of the radionuclide allows tailoring the properties of the labeled protein to the application required. Until the event of positron emission tomography the spectrum of nuclides used to visualize cellular and biochemical processes was largely restricted to iodine isotopes and 99m-technetium. Today, several nuclides such as 18-fluorine, 68-gallium and 86-yttrium have fundamentally extended the possibilities of tracer design and in turn caused the need for the development of chemical methods for their conjugation.

Alpha v beta 6 (αvβ6) integrin is overexpressed by several carcinomas, and thus considered as target for diagnostic imaging and anti-cancer therapies. Recently, we presented SFITGv6 as a novel potential tracer for imaging and targeted therapy of αvβ6 integin-positive carcinomas. Here, we analyzed affinity and specificity of five native αvβ6 integrin-specific binders in comparison to SFITGv6. Methods: Sunflower trypsin inhibitor (SFTI)-1-based peptides containing arginine-glycine-aspartic acid (RGD) motif-spanning octamers of fibronectin (SFFN1), tenascin C (SFTNC), vitronectin (SFVTN), and latency-associated peptides (LAP) 1 (SFLAP1), and 3 (SFLAP3) were synthesized and their binding potential to αvβ6 integrin-expressing head and neck squamous cell carcinoma (HNSCC) cell lines was evaluated. Subsequently, stability, affinity and specificity were assessed in vitro using radio-high pressure liquid chromatography, surface plasmon resonance (SPR) assay, and binding experiments including competition, kinetics, internalization, and efflux. αvβ6 integrin binding specificity was further evaluated by peptide histochemistry. Finally, in vivo binding properties were assessed using small-animal positron emission tomography (PET) imaging and biodistribution experiments in HNSCC-bearing mice and ⁶⁸Ga-DOTA-SFLAP3 was applied for diagnostic positron emission tomography/computed tomography (PET/CT) of a HNSCC patient. Results: Comparing the newly designed peptides significant binding (>20%) to several HNSCC cell lines (HNO97, HNO399, and HNO223) and a fast internalization of up to 60% and 70% was observed for SFLAP3 (GRGDLGRL) and SFITGv6 (FRGDLMQL). In contrast, the other peptides displayed moderate (SFLAP1: 4.1-12.1%) to marginal binding (SFFN1, SFTNC and SFVTN: <1%) and were, therefore, excluded from further analysis. Notably, SFLAP3 exhibited improved affinity for αvβ6 integrin (IC50mean = 3.5 nM; KD = 7.4). Moreover, small- animal positron emission tomography (PET) imaging and biodistribution studies of HNSCC xenograft mice revealed an increased tumor-specific accumulation 30-60 min after injection of ¹⁷⁷Lu-labeled and/or ⁶⁸Ga-DOTA-labeled SFLAP3. Peptide staining further demonstrated binding specificity of SFLAP3 to HNSCC tumor cells. Finally, PET/CT scan of an HNSCC patient showed specific SFLAP3 accumulation in the primary tumor lesion (standardized uptake values (SUV)max = 5.1) and in a corresponding lymph node metastases (SUV_max = 4.1). Conclusion: SFLAP3 represents a promising tracer for prognostic assessment, diagnostic imaging and possibly targeted therapy of αvβ6 integrin-expressing tumors.