Transitioning the Molecular Tumor Board from Proof of Concept to Clinical Routine: A German Single-Center Analysis.

Molecular precision oncology faces two major challenges: first, to identify relevant and actionable molecular variants in a rapidly changing field and second, to provide access to a broad patient population. Here, we report a four-year experience of the Molecular Tumor Board (MTB) of the Comprehensive Cancer Center Freiburg (Germany) including workflows and process optimizations. This retrospective single-center study includes data on 488 patients enrolled in the MTB from February 2015 through December 2018. Recommendations include individual molecular diagnostics, molecular stratified therapies, assessment of treatment adherence and patient outcomes including overall survival. The majority of MTB patients presented with stage IV oncologic malignancies (90.6%) and underwent an average of 2.1 previous lines of therapy. Individual diagnostic recommendations were given to 487 patients (99.8%). A treatment recommendation was given in 264 of all cases (54.1%) which included a molecularly matched treatment in 212 patients (43.4%). The 264 treatment recommendations were implemented in 76 patients (28.8%). Stable disease was observed in 19 patients (25.0%), 17 had partial response (22.4%) and five showed a complete remission (6.6%). An objective response was achieved in 28.9% of cases with implemented recommendations and for 4.5% of the total population (22 of 488 patients). By optimizing the MTB workflow, case-discussions per session increased significantly while treatment adherence and outcome remained stable over time. Our data demonstrate the feasibility and effectiveness of molecular-guided personalized therapy for cancer patients in a clinical routine setting showing a low but robust and durable disease control rate over time.

Differences in isoprenoid-mediated energy dissipation pathways between coastal and interior Douglas-fir seedlings in response to drought.

Plants have evolved energy dissipation pathways to reduce photooxidative damage under drought when photosynthesis is hampered. Non-volatile and volatile isoprenoids are involved in non-photochemical quenching of excess light energy and scavenging of reactive oxygen species. A better understanding of trees' ability to cope with and withstand drought stress will contribute to mitigate the negative effects of prolonged drought periods expected under future climate conditions. Therefore we investigated if Douglas-fir (Pseudotsuga menziesii(Mirb.)) provenances from habitats with contrasting water availability reveal intraspecific variation in isoprenoid-mediated energy dissipation pathways. In a controlled drought experiment with 1-year-old seedlings of an interior and a coastal Douglas-fir provenance, we assessed the photosynthetic capacity, pool sizes of non-volatile isoprenoids associated with the photosynthetic apparatus, as well as pool sizes and emission of volatile isoprenoids. We observed variation in the amount and composition of non-volatile and volatile isoprenoids among provenances, which could be linked to variation in photosynthetic capacity under drought. The coastal provenance exhibited an enhanced biosynthesis and emission of volatile isoprenoids, which is likely sustained by generally higher assimilation rates under drought. In contrast, the interior provenance showed an enhanced photoprotection of the photosynthetic apparatus by generally higher amounts of non-volatile isoprenoids and increased amounts of xanthophyll cycle pigments under drought. Our results demonstrate that there is intraspecific variation in isoprenoid-mediated energy dissipation pathways among Douglas-fir provenances, which may be important traits when selecting provenances suitable to grow under future climate conditions.

Foliar nitrogen metabolism of adult Douglas-fir trees is affected by soil water availability and varies little among provenances.

The coniferous forest tree Douglas-fir (Pseudotsuga menziesii) is native to the pacific North America, and is increasingly planted in temperate regions worldwide. Nitrogen (N) metabolism is of great importance for growth, resistance and resilience of trees. In the present study, foliar N metabolism of adult trees of three coastal and one interior provenance of Douglas-fir grown at two common gardens in southwestern Germany (Wiesloch, W; Schluchsee, S) were characterized in two subsequent years. Both the native North American habitats of the seed sources and the common garden sites in Germany differ in climate conditions. Total and mineral soil N as well as soil water content were higher in S compared to W. We hypothesized that i) provenances differ constitutively in N pool sizes and composition, ii) N pools are affected by environmental conditions, and iii) that effects of environmental factors on N pools differ among interior and coastal provenances. Soil water content strongly affected the concentrations of total N, soluble protein, total amino acids (TAA), arginine and glutamate. Foliar concentrations of total N, soluble protein, structural N and TAA of trees grown at W were much higher than in trees at S. Provenance effects were small but significant for total N and soluble protein content (interior provenance showed lowest concentrations), as well as arginine, asparagine and glutamate. Our data suggest that needle N status of adult Douglas-fir is independent from soil N availability and that low soil water availability induces a re-allocation of N from structural N to metabolic N pools. Small provenance effects on N pools suggest that local adaptation of Douglas-fir is not dominated by N conditions at the native habitats.

Drought effects on root and needle terpenoid content of a coastal and an interior Douglas fir provenance.

Douglas fir (Pseudotsuga menziesii) is a conifer species that stores large amounts of terpenoids, mainly monoterpenoids in resin ducts of various tissues. The effects of drought on stored leaf terpenoid concentrations in trees are scarcely studied and published data are partially controversial, since reduced, unaffected or elevated terpenoid contents due to drought have been reported. Even less is known on the effect of drought on root terpenoids. In the present work, we investigated the effect of reduced water availability on the terpenoid content in roots and needles of Douglas fir seedlings. Two contrasting Douglas fir provenances were studied: an interior provenance (var. glauca) with assumed higher drought resistance, and a coastal provenance (var. menziesii) with assumed lower drought resistance. We tested the hypothesis that both provenances show specific patterns of stored terpenoids and that the patterns will change in response to drought in both, needles and roots. We further expected stronger changes in the less drought tolerant coastal provenance. For this purpose, we performed an experiment under controlled conditions, in which the trees were exposed to moderate and severe drought stress. According to our expectations, the study revealed clear provenance-specific terpenoid patterns in needles. However, such patterns were not detected in the roots. Drought slightly increased the needle terpenoid contents of the coastal but not of the interior provenance. We also observed increased terpenoid abundance mainly in roots of the moderately stressed coastal provenance. Overall, from the observed provenance-specific reactions with increased terpenoid levels in trees of the coastal origin in response to drought, we conclude on functions of terpenoids for abiotic stress tolerance that might be fulfilled by other, constitutively expressed mechanisms in drought-adapted interior provenances.

Variation in short-term and long-term responses of photosynthesis and isoprenoid-mediated photoprotection to soil water availability in four Douglas-fir provenances.

For long-lived forest tree species, the understanding of intraspecific variation among populations and their response to water availability can reveal their ability to cope with and adapt to climate change. Dissipation of excess excitation energy, mediated by photoprotective isoprenoids, is an important defense mechanism against drought and high light when photosynthesis is hampered. We used 50-year-old Douglas-fir trees of four provenances at two common garden experiments to characterize provenance-specific variation in photosynthesis and photoprotective mechanisms mediated by essential and non-essential isoprenoids in response to soil water availability and solar radiation. All provenances revealed uniform photoprotective responses to high solar radiation, including increased de-epoxidation of photoprotective xanthophyll cycle pigments and enhanced emission of volatile monoterpenes. In contrast, we observed differences between provenances in response to drought, where provenances sustaining higher CO2 assimilation rates also revealed increased water-use efficiency, carotenoid-chlorophyll ratios, pools of xanthophyll cycle pigments, ß-carotene and stored monoterpenes. Our results demonstrate that local adaptation to contrasting habitats affected chlorophyll-carotenoid ratios, pool sizes of photoprotective xanthophylls, ß-carotene, and stored volatile isoprenoids. We conclude that intraspecific variation in isoprenoid-mediated photoprotective mechanisms contributes to the adaptive potential of Douglas-fir provenances to climate change.

A coastal and an interior Douglas fir provenance exhibit different metabolic strategies to deal with drought stress.

Drought is a major environmental stress affecting growth and vitality of forest ecosystems. In the present study, foliar nitrogen (N) and carbon (C) metabolism of two Douglas fir (Pseudotsuga menziesii) provenances with assumed different drought tolerance were investigated. We worked with 1-year-old seedlings of the interior provenance Fehr Lake (FEHR) originating from a dry environment and the coastal provenance Snoqualmie (SNO) from a more humid origin. Total C and N, structural N and the concentrations of soluble protein, total amino acids (TAAs) and individual amino acids as well as the relative abundance of polar, low-molecular-weight metabolites including antioxidants were determined in current-year needles exposed either to 42 days of drought or to 42 days drought plus 14 days of rewatering. The seedlings reacted in a provenance-specific manner to drought stress. Coastal provenance SNO showed considerably increased contents of TAAs, which were caused by increased abundance of the quantitatively most important amino acids arginine, ornithine and lysine. Additionally, the polyamine putrescine accumulated exclusively in drought-stressed trees of this provenance. In contrast, the interior provenance FEHR showed the opposite response, i.e., drastically reduced concentrations of these amino acids. However, FEHR showed considerably increased contents of pyruvate-derived and aromatic amino acids, and also higher drought-induced levels of the antioxidants ascorbate and α-tocopherol. In response to drought, both provenances produced large amounts of carbohydrates, such as glucose and fructose, most likely as osmolytes that can readily be metabolized for protection against osmotic stress. We conclude that FEHR and SNO cope with drought stress in a provenance-specific manner: the coastal provenance SNO was mainly synthesizing N-based osmolytes, a reaction not observed in the interior provenance FEHR; instead, the latter increased the levels of scavengers of reactive oxygen species. Our results underline the importance of provenance-specific reactions to abiotic stress.