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BACKGROUND BRCA1 and BRCA2 genes represent the most investigated breast and ovarian cancer predisposition genes. Ten cases of pathogenic de novo BRCA1 variations and six cases of pathogenic de novo BRCA2 variation have been reported at present. Here, we report a new case of a de novo BRCA1 gene mutation. CASE PRESENTATION A 30-year-old woman with no health issues and no family history for hereditary breast and ovarian cancer was diagnosed with a hormone receptor positive/HER2 negative invasive breast cancer. Genetic testing revealed a pathogenic variant in BRCA1 (c.4065_4068delTCAA) which was not found in her parents or sister. CONCLUSION We report a new case of de novo BRCA1 mutation, confirmed by repeated germline testing of the index patient and her parents. The published BRCA1/2 de novo mutation rate is low. This is probably due - in part - to the strict testing criteria.

The Second International Symposium on Cellular and Organismal Stress Responses took place virtually on September 8–9, 2022. This meeting was supported by the Cell Stress Society International (CSSI) and organized by Patricija Van Oosten-Hawle and Andrew Truman (University of North Carolina at Charlotte, USA) and Mehdi Mollapour (SUNY Upstate Medical University, USA). The goal of this symposium was to continue the theme from the initial meeting in 2020 by providing a platform for established researchers, new investigators, postdoctoral fellows, and students to present and exchange ideas on various topics on cellular stress and chaperones. We will summarize the highlights of the meeting here and recognize those that received recognition from the CSSI.

Blommersia bara sp. nov. Figures 4, 8 Identity. This species was considered Mantidactylus wittei by Guibé (1974) and Blommers-Schl̂sser & Blanc (1991) for specimens from Ankarafantsika, and called Mantidactylus cf. wittei by Glos (2003), Blommersia sp. aff. wittei “Isalo” by Glaw & Vences (2007), Blommersia sp. aff. wittei by Mercurio et al. (2008), Blommersia sp. (aff. wittei) by Bora et al. (2010), Blommersia sp. 5 by Vieites et al. (2009) and Wollenberg et al. (2011), Blommersia sp. Ca5 by Perl et al. (2014), Penny et al. (2016) and Glaw et al. (2019), Blommersia sp. Ca05 by Penny et al. (2017), and Blommersia sp. aff. wittei Ca 05 “Isalo” by Cocca et al. (2018). Holotype. ZSM 31/2004 (field number FGZC 47), an adult male, collected by F. Glaw, M. Puente, M. Thomas & R. Randrianiaina at a stream near Ranohira, Isalo Massif (22.5856°S, 45.3997°E, 813 a.s.l.), southwestern Madagascar, on 21 January 2004. Paratypes. A total of 30 specimens, all from western Madagascar. ZSM 22/2004 (FGZC 34) and ZSM 24/2004 (FGZC 38) two adult males with the same collecting data as the holotype; ZSM 706/2001 (FGMV 2001.279), adult male, collected by M. Vences, D.R. Vieites, G. Garcia, V. Raherisoa, & A. Rasoamamonjinirina at Ampijoroa, Ankarafantsika National Park (approximately 16.3°S, 46.82°E) on 24 February 2001; ZSM 246/2003 and ZSM 247/2003, two adult males, collected by J. Glos at Kirindy Forest (site “G2”) in January 2001; ZSM 13/2006 (FGZC 691), ZSM 30/2006 (FGZC 727), ZSM 52/2006 (FGZC 779), and ZSM 68/2006 (FGZC 809), two adult females and two adult males, collected by F. Glaw, J. K̂hler, P. Bora & H. Enting at Tsingy de Bemaraha National Park, Andranopasazy, “Camp 1” (18.7086°S, 44.7189°E, 146 m a.s.l.), on 17, 19, and 24 March 2006, respectively; ZSM 2284/2007 (ZCMV 5801) and ZSM 2285/2007 (ZCMV 5804), one adult male and one adult female, collected by M. Vences and collaborators at Isalo National Park, on 17 February 2007; ZSM 2281/2007 (ZCMV 5625), ZSM 2282/2007 (ZCMV 5626), ZSM 2283/2007 (ZCMV 5643), and ZSM 2320/2007 (ZCMV 5621), collected by L. du Preez, C. Weldon & L. Raharivololoniaina at Ankarafantsika, on 9 February 2007; ZSM 3222/2012 (ZCMV 14143), adult male collected by A. Rakotoarison, J. Erens & E. Rajeriarison at Mariarano (15.4978°S, 46.6943°E, 11 m a.s.l.), on 28 December 2012; MRSN A2957 and A2958 (FAZC 11807–11808), two males, collected by F. Andreone, G. Aprea & V. Mercurio in the Isalo Massif, Andohasahenina (22.8333°S, 45.1880°E, 876 m a.s.l.) on 15 January 2004; MRSN A5349 (FAZC 12550), female, and MRSN A5350 (FAZC 12551), male, collected by F. Andreone, F. Mattioli & V. Mercurio in the Isalo Massif, Zahavola (22.6215°S, 45.3587°E, 881 m a.s.l.), on 17 November 2004; MRSN A5353 and A5354 (FAZC 12968 and 12970), two males, collected by F. Andreone, F. Mattioli & V. Mercurio in the Isalo Massif, Sakamalio (22.4348°S, 45.2552°E, 726 m a.s.l.) on 16 December 2004; MRSN A5351 (FAZC 12591), female, collected by F. Andreone, F. Mattioli & V. Mercurio in the Isalo Massif, Andriamanero (22.3672°S, 45.3920°E, 663 m a.s.l.), on 20 November 2004; as well as seven specimens deposited in the UADBA collection (uncatalogued but accessible via field tags): UADBA-ZCMV 5620, 5641, 5642, 5644, four specimens collected by L. du Preez, C. Weldon & L. Raharivololoniaina at Ankarafantsika, on 9 February 2007; and UADBA-ZCMV 5800, 5802, 5803, three specimens collected by collected by M. Vences and collaborators at Isalo National Park, on 17 February 2007. Diagnosis. A species of the genus Blommersia in the subfamily Mantellinae of the family Mantellidae based on presence of intercalary elements between penultimate and ultimate phalanges of fingers and toes (verified by external examination), occurrence in Madagascar, relatively small body size (male SVL From other species of Blommersia, the new species is mainly distinguished as follows: from B. angolafa by the presence of vomerine teeth in many individuals (vs. absence) and a totally different color pattern without whitish spots on the flanks and on the finger- and toe-tips (vs. presence); from B. dejongi by a femoral gland placed centrally on the thigh (vs. distally); from B. blommersae by having lateral metatarsalia separated by webbing (vs. connected by dense tissue); from B. domerguei by having lateral metatarsalia separated by webbing (vs. connected by dense tissue), by the presence of vomerine teeth in many individuals (vs. absence), a larger body size (SVL> 20 mm vs. B. dupreezi by having lateral metatarsalia separated by webbing (vs. connected by dense tissue) and absence of distinct black lateral stripe (vs. presence); from B. galani by presence of vomerine teeth in many individuals (vs. absence); from B. grandisonae by presence of vomerine teeth in many individuals (vs. absence) and a largely different color pattern (but see discussion in Vences et al. 2023 on the identity of B. grandisonae); from B. kely by having lateral metatarsalia separated by webbing (vs. connected by dense tissue), presence of vomerine teeth in many individuals (vs. absence), and larger body size (SVL> 20 mm vs. B. nataliae by ovoid femoral glands in small distance to each other (vs. more rounded, distant glands) and absence of a dark face mask (vs. presence); from B. sarotra by having lateral metatarsalia separated by webbing (vs. connected by dense tissue); from B. transmarina by a narrower head (male HW/SVL ratio 0.29‒0.34 vs. 0.34–0.36) and somewhat shorter hands (male HAL/SVL ratio 0.29–0.33 vs. 0.30–0.35); from B. variabilis by having lateral metatarsalia separated by webbing (vs. connected by dense tissue in most individuals) and probably a smaller distance between femoral gland. Furthermore, differentiated from all these species by differences in advertisement calls and genetic distances>5% in the analyzed mitochondrial 16S rRNA gene fragment of these two species. It differs from B. wittei, however, in a substantial divergence of mitochondrial genes (5.0‒8.2% uncorrected pairwise distance in 16S) and lack of haplotype sharing in RAG1 and SACS, and in advertisement calls lacking distinct pulses vs. being clearly pulsed in B. wittei (except in the Sambava population whose taxonomic status is in need of revision). Description of the holotype. Adult male specimen with distinct femoral glands, in a good state of preservation (Fig. 8). Tongue removed as tissue samples for molecular analysis. SVL 18.5 mm, for further measurements see Table 1. Body slender; head longer than wide, of same width as body; snout rounded in lateral view, obtusely pointed in dorsal and ventral views; nostrils directed laterally, protuberant, nearer to snout tip than to eye; canthus rostralis rounded; loreal region slightly concave; tympanum distinct, round, its diameter 63% of eye diameter; supratympanic fold distinct, curved above tympanum where it follows the tympanum outline; tongue absent, its shape therefore not ascertainable; vomerine teeth not visible; choanae small, round, located toward the front of the palate; maxillary teeth present.Arms slender, subarticular tubercles present, single; fingers without webbing; relative length of fingers 1 After 19 years in preservative (Fig. 8), the dorsum is rather uniformly brownish, with a weakly recognizable dark brownish inverted chevron, a faint small dark transverse bar on the forehead anterior to the eyes, and a very weakly recognizable dark interorbital bar. A dark stripe runs from the nostril to the eye and continues broader posterior to the eye, encompassing the entire tympanic region underneath the supratympanic fold. Underneath these brown elements, a weakly expressed light frenal stripe is visible from the snout tip to the forelimb insertions. Fore- and hindlimbs dorsally with distinct and sharply delimited brown crossbars (three crossbars on thigh and four on shank). Ventral side uniformly beige (unpigmented), throat being a bit lighter than chest, belly and limbs. In life, color was very similar but somewhat more contrasted (Fig. 4A); whitish color was present on the throat; femoral glands granules were somewhat yellowish (Fig. 4B). Variation. ZSM 52/2006 exhibits a broad middorsal light band. Vomerine teeth are clearly visible in paratype ZSM 2281/2007 from Ankarafantsika, also recognizable (although weakly expressed) in the paratypes from Bemaraha. We observed substantial differences in body size among populations, with individuals from the type locality Isalo being distinctly smaller than those from other sites. According to available data (Supplementary Table 2; available at DOI 10.5281/zenodo.8049142) male SVL across all populations ranges between 18.2–25.7 mm and female SVL between 22.0– 26.4 mm. Etymology. Named after the Bara people, the ethnic group living in the area of Madagascar that includes the type locality of the new species, the Isalo Massif. The name is used as a noun in apposition. Natural history. A relatively common species in Madagascar, living in areas of a certain humidity such as streams or swamps, including areas of secondary vegetation. During the rainy season, males call during day and night from the low vegetation of these water bodies. In Isalo this species is commonly found on the ground of gallery forests (e.g., Andriamanero, Sakamalio), along temporary rivers and in Pandanus swamps in the open savannah (e.g., Ilakaka, Zahavola). Tadpoles develop in temporary ponds and individuals of this species have been observed in egg-guarding behaviour both in Ankarafantsika (Fig. 4C) and Isalo (Mercurio et al. (2008). In the Sahamalaza Peninsula, this species seems to be abundant and it is found along streams and ponds in intact forested areas as well as in paddy fields in cleared areas. In Bemaraha, specimens were found in the leaf litter. In Kirindy, Glos (2003) found the species in the largest out of 200 ponds studied, and observed males calling at night from a bush at the water edge between 1–4 m above ground. Vocalizations: Advertisement calls recorded on 29 January 1994 at Isalo (air temperature 21.5°C) consist of a single short pulsatile note repeated in call series at rather regular intervals within series (Fig. 6). Each call exhibits some irregular amplitude modulation, sometimes with 2–4 peaks recognizable within each call, with the second peak usually having the highest energy. Numerical parameters of 23 analyzed calls of one male are as follows: call duration (= note duration) 13–18 ms (15.2 ± 1.6 ms); inter-call intervals within regular call series 69–78 ms (74.8 ± 3.0 ms); duration of regular call series 2034 ms (n = 1); call rate within call series approximately 670 calls/minute; dominant frequency 5813–6115 Hz (5968 ± 119 Hz); second dominant frequency peak around 3000 Hz; prevalent bandwidth 2600–9500 Hz. Advertisement calls recorded on 24 February 2001 at Ankarafantsika (air temperature 28°C) consist of a single short pulsatile note repeated in call series at regular intervals within series (Fig. 6). In some calls (= notes) two barely separated peaks of amplitude are recognizable, but call energy is distributed rather equally along the first two thirds of the call’s duration, then dropping rapidly towards its end. Numerical parameters of 18 analyzed calls of one male are as follows: call duration (= note duration) 19–28 ms (23.2 ± 2.5 ms); inter-call intervals within regular call series 94–107 ms (97.3 ± 4.8 ms); duration of regular call series 2078 ms (n = 1); call rate within call series approximately 500 calls/minute; dominant frequency 4694–4758 Hz (4735 ± 27 Hz); second frequency peak around 2370 Hz; prevalent bandwidth 1900–9700 Hz. Distribution. Known from Isalo in the South-West up to Sahamalaza Peninsula in the North-West (see Fig. 2 for genetically confirmed sites). The precise contact zone with B. wittei has not yet been identified but can be expected to be located between Sahamalaza and Ambanja.

Blommersia wittei (Guibé, 1974) Figures 3, 8 Identity and type material. Clarifying the taxonomy of the B. wittei complex requires first ascertaining the identity of B. wittei sensu stricto. The species was described by Guibé (1974) as Mantidactylus wittei, and type specimens are deposited in the Paris museum (acronym MNHN, in earlier times written MNHNP), with the following verbatim information in the original description: “ Holotype: no 1953-60 MNHN Paris. Environs d’Ambanja. J. Guibé (11- 1951). Paratypes: nos 1953-60 A à 1953-60 L MNHN Paris. Même provenance. Nos A.682 à A.684: Ampijora , station forestière à 45 km de Marovaoy (Ouest). Nos 1973-944 à 1973-951 MNHN Paris: forêt d’Ankarafantsika. Ch. P. Blanc (11-1973). ” Due to some imprecise information in the handwritten MNHN catalogue and extensive re-labelling of several of these specimens, there has been confusion on the type series in subsequent publications and in the current digital MNHN catalogue. Frost (2023) correctly states that the holotype is MNHNP 1953.60, by original designation, but merges two separate sites in the type locality account: “ ‘ Environs d’Ambanja’, Forest Ankarafantsika, Madagascar. ” In contrast, the digital type catalogue of the MNHN (downloaded from gbif.org in 2022) as well as the MNHN online catalogue (https://science.mnhn.fr/institution/mnhn/collection/ra/; accessed 11 February 2023) state that the holotype originated from “ Vallée de la Tsiribihina ” and furthermore provide the following list of paratypes: MNHN 1973.944–1973.951 (8 specimens) from Ankarafantsika, and 1993.682–1993.684 (3 specimens) from Ampijoroa. Vences et al. (2010) provided measurements of part of the type series of B. wittei, purportedly all from Ambanja: the holotype MNHN 1953.60, and the following paratypes: MNHN 1991.2529–1991.2533 (5 specimens; previously labelled MNHN 1953.60 A to E) and MNHN 1991.2536–1991.2539 (4 specimens, previously labelled MNHN 1953.60 H to K). After inspecting the original (handwritten) MNHN catalogues (partly available from https://science.mnhn.fr/ catalogues; accessed 11 February 2023) we conclude that: (1) The holotype of Mantidactylus wittei Guibé, 1974 (Fig. 8) unambiguously is the specimen MNHN 1953.60 for which morphometric measurements were provided by Vences et al. (2010) and whose locality, according to the original description and original catalogue entry is “ Environs d’Ambanja”, with the addition in the catalogue: “Cacaoyéres” (= cacao trees, thus indicating the specimen was collected in a cacao plantation). (2) A series of 11 paratypes (originally labelled MNHN 1953.60 A to 1953 K, currently re-labelled as MNHN 1991.2529–1991.2539) originates from the same locality as the holotype. (3) A series of 8 paratypes (MNHN 1973.944–1973.951) originate from Ankarafantsika, (4) A series of 3 paratypes (MNHN 1993.682–1993.684) originate from Ampijoroa, which represents a forestry station (currently local headquarters of Madagascar National Parks) within the Ankarafantsika forest. As explained in the Taxonomic conclusion account above, based on the provenance of the holotype from Ambanja, we continue assigning the name B. wittei to the genetic lineage occurring across much of northern Madagascar and collected by us at Ambanja and nearby localities. The paratype series is however mixed: specimens from Ankarafantsika (MNHN 1973.944–1973.951) and Ampijoroa (MNHN 1993.682–1993.684) are here assigned to B. sp. Ca 5. Only the 11 paratypes from the type locality are likely conspecific with the holotype. Morphology. Measurements of the type series of B. wittei have been published by Vences et al. (2010) and measurements of additional specimens by Pabijan et al. (2011). Measurements of further specimens are included in Table 1 herein.A full set of measurements including those from the previous publications is given as Supplementary Table 2 (available from the Zenodo repository under DOI 10.5281/zenodo.8049142). Based on these data, male SVL is 21.0–27.0 mm and female SVL is 20.7–25.0 mm. In the specimens examined for the present study (listed in Table 1), vomerine teeth are usually recognizable (clearly visible in specimens from the type locality Ambanja) but weakly expressed in several specimens from Montagne d’Ambre. In ZSM 563/2000 from Sambava, vomerine teeth are weakly recognizable, and this specimen apparently has more developed webbing than many other individuals examined. Vocalizations.Advertisement calls recorded in February 1991 at the type locality Ambanja (recording temperature not taken) consist of a single short pulsed note repeated in call series at regular intervals and very fast succession (Fig. 5). Recording quality is poor and the detailed call structure is probably partly masked by background noises. However, each call (= note) seems to contain two pulse groups which are separated from each other, with the first pulse group being of lower amplitude. Pulse structure in not very obvious in the recording and most pulses appear basally fused. Maximum call energy is distributed towards the middle of the call’s duration. Numerical parameters of 16 analyzed calls of one male are as follows: call duration (= note duration) 62–77 ms (66.8 ± 5.3 ms); inter-call intervals within regular call series 15–31 ms (18.6 ± 5.6 ms); pulses/note 9–18 (13.8 ± 3.3); duration of regular call series 1275 ms (n = 1); call rate within call series approximately 740 calls/minute; dominant frequency 4694–4886 Hz (4797 ± 82 Hz); second frequency peak around 2500 Hz; prevalent bandwidth 1800–6000 Hz. Advertisement calls recorded on 7 February 1992 from north of Andoany, Nosy Be (air temperature 25°C), consist of a single short pulsed note repeated in short call series at regular intervals and very fast succession (Fig. 5). Each call (= note) exhibits two pulse groups which are clearly separated from each other, with the first pulse group being of lower amplitude. Pulses are partly fused, but countable. Maximum call energy is distributed in the middle of the call, namely the beginning of the second pulse group. Numerical parameters of 10 analyzed calls of one male are as follows: call duration (= note duration) 65–83 ms (74.8 ± 6.3 ms); inter-call intervals within regular call series 16–32 ms (21.5 ± 5.6 ms); pulses/note 14–19 (16.3 ± 1.9); duration of regular call series 490 and 500 ms (n = 2); call rate within call series approximately 640 calls/minute; dominant frequency 4823–5240 Hz (5033 ± 163 Hz); second frequency peak around 2500 Hz; prevalent bandwidth 2000–8200 Hz. Advertisement calls recorded on 15 March 1994 on Montagne d’Ambre (air temperature 22°C), consist of a single short, distinctly pulsed note repeated in call series at regular intervals and very fast succession (Fig. 5). Each call (= note) exhibits two pulse groups which are clearly separated from each other, with the first pulse group being of lower amplitude. Pulses are barely fused and rather distinctly separated. Maximum call energy is distributed in the middle of the call, namely the beginning of the second pulse group. Numerical parameters of 19 analyzed calls of one individual are as follows: call duration (= note duration) 59–73 ms (65.4 ± 4.2 ms); inter-call intervals within regular call series 22–39 ms (29.6 ± 7.1 ms); pulses/note 7–11 (8.9 ± 1.3); duration of regular call series 1821 ms (n = 1); call rate within call series approximately 600 calls/minute; dominant frequency 4489–4597 Hz (4542 ± 41 Hz); prevalent bandwidth 1800–7500 Hz. Advertisement calls recorded from specimen ZSM 50/2018 (MSZC 521) on 25 December 2017 on Montagne d’Ambre (air temperature ca 19°C), consist of a single short, distinctly pulsed note repeated in call series at regular intervals and very fast succession (Fig. 5). Each call (= note) exhibits clearly separated pulses. Intervals between pulses are somewhat irregular, in some calls resulting in two pulse groups separated by a larger interval. Maximum call energy is evident in the middle of the call. Numerical parameters of 16 analyzed calls of one male are as follows: call duration (= note duration) 68–88 ms (78.2 ± 5.7 ms); inter-call intervals within regular call series 28–50 ms (41.4 ± 4.2 ms); pulses/note 7–11 (9.0 ± 1.3); duration of regular call series 1590 ms (n = 1); call rate within call series approximately 500 calls/minute; dominant frequency 5066–5232 Hz (5137 ± 80 Hz); prevalent bandwidth 2000–10000 Hz. Advertisement calls recorded on 20 March 2000 in Sambava (air temperature 24.7°C), here tentatively allocated to B. wittei, consist of a single, short, pulsed note repeated in call series at regular intervals within series. Calls (= notes) exhibit a pulsed structure, but most pulses are largely fused, resulting in an irregular pulse pattern and varying number of countable pulses (Fig. 7). Maximum call energy is distributed among the first two thirds of the call’s duration. Numerical parameters of 22 analyzed calls of one male are as follows: call duration (= note duration) 27– 59 ms (39.4 ± 11.8 ms); inter-call intervals within regular call series 54–76 ms (67.9 ± 7.7 ms); pulses/note 2–6 (4.1 ± 1.0); duration of regular call series 2209 ms (n = 1); call rate within call series approximately 600 calls/minute; dominant frequency 5316–5598 Hz (5440 ± 145 Hz); second frequency peak around 2600 Hz; prevalent bandwidth 2100–8600 Hz.

Blommersia sp. Ca12 Identity. This candidate species name refers to a new lineage discovered in the course of this study and found only at two sites along the Sambirano river (Maevatanana and Antsirasira). Morphology. Unknown. One voucher specimen from Antsirasira (ZSM 589/2001; see Table 1) could not be genotyped as no tissue sample for molecular analysis was taken in the field from this specimen. This specimen is an adult male with rather indistinct femoral glands and weakly expressed vomerine teeth. Photos in life are not available. Vocalizations: Advertisement calls were recorded on 31 January 2001 at Antsirasira (air temperature 26°C), but call vouchers were not collected and calls are therefore not assignable to B. sp. Ca12 with full reliability. These calls consist of a single, short, pulsed note repeated in call series at slightly irregular intervals within series (Fig. 7). Calls (= notes) generally exhibit a pulse structure, although some pulses are not clearly separated but rather largely fused, resulting in an irregular pulse pattern and varying number of countable pulses. Numerical parameters of 10 analyzed calls of one male are as follows: call duration (= note duration) 34–66 ms (47.8 ± 10.8 ms); inter-call intervals within regular call series 48–72 ms (61.4 ± 10.2 ms); pulses/note 3–10 (6.9 ± 2.3); duration of regular call series 1434 and 995 ms (n = 2); call rate within call series varied between 500–580 calls/minute; dominant frequency 5316–5598 Hz (5440 ± 145 Hz); second frequency peak around 2850 Hz; prevalent bandwidth 2200–9000 Hz.

Vences, Miguel, Multzsch, Malte, Köhler, Jörn, Crottini, Angelica, Andreone, Franco, Rakotoarison, Andolalao, Scherz, Mark D., Glaw, Frank (2023): Integrative revision of the Blommersia wittei complex, with description of a new species of frog from western and north-western Madagascar. Zootaxa 5319 (2): 178-198, DOI: 10.11646/zootaxa.5319.2.2, URL: http://dx.doi.org/10.11646/zootaxa.5319.2.2

Guibemantis (Pandanusicola) pulcherrimus sp. nov. Figs. 7‒9 Holotype. ZSM 518/2009 (field number ZCMV 11217), adult female, collected by M. Vences, D.R. Vieites, F.M. Ratsoavina, R.D. Randrianiaina, E. Rajeriarison, T. Rajoafiarison and J.L. Patton on 23 June 2009 near a site locally named Hevirina, western slope of the Makira Massif, Analanjirofo Region, Madagascar (GPS coordinates not taken, but coordinates of Hevirina are 15.4490°S, 49.1119°E, 1093 m a.s.l.). Paratypes. Eight specimens. ZSM 519/2009 (ZCMV 11249) and ZSM 520/2009 (ZCMV 11264), two adult females, collected by M. Vences, D.R. Vieites, F.M. Ratsoavina, R.D. Randrianiaina, E. Rajeriarison, T. Rajoafiarison and J.L. Patton on 23–24 June 2009 at a campsite near the source of Fotsialanana river, western slope of the Makira Massif, Analanjirofo Region, Madagascar (coordinates 15.46675°S, 49.128889°E, 1067 m a.s.l.); ZSM 1754/2010 (ZCMV 12505), adult female, collected by M. Vences, D.R. Vieites, R.D. Randrianiaina, F.M. Ratsoavina, S. Rasamison, A. Rakotoarison, E. Rajeriarison and T. Rajoafiarison on 27 June 2010 at Bemanevika, campsite Atsirakala, Sofia Region, Madagascar (coordinates 14.43061°S, 48.60179°E, 1466 m a.s.l.); as well as five specimens (field numbers ZCMV 11225, ZCMV 11226, ZCMV 11228, ZCMV 11229), all collected in June 2009 by by M. Vences, D.R. Vieites, R.D. Randrianiaina, F.M. Ratsoavina, S. Rasamison, A. Rakotoarison, E. Rajeriarison and T. Rajoafiarison on the western slope the Makira Massif, not examined morphologically but included in the 16S tree (Fig. 1), integrated but still uncatalogued at the UADBA (MZBA) collection (individually identifiable via their field number tags). Diagnosis. Assigned to the subgenus Pandanusicola of the genus Guibemantis based on its small body size, phytotelm-dwelling habitats (in Pandanus plants), near-absent webbing between toes, connected lateral metatarsalia, the presence of both inner and outer metatarsal tubercles, intercalary elements between ultimate and penultimate phalanges of fingers and toes (verified by external examination), and molecular phylogeny. The new species is characterized by the unique combination of the following characters: (1) medium size with SVL 25‒29 mm in females, (2) bright green dorsal color with translucent shade, with dense fine purplish-brown spots, (3) distinct brown rostral stripe, (4) absence of white or yellow spots in the inguinal region, (5) absence of webbing on hand and only traces of web at foot, (6) vomerine teeth rudimentary. The new species is most similar to its sister species G. pulcher, which differs (despite some overlap) by fewer (12–107, mean 42; vs. 47–223, mean 103) and larger (diameter often up to 0.8 mm vs. 0.4 mm) purplish-brown dorsal spots, and a larger and denser purplish-brown band on the flanks which often is adorned centrally by silvery whitish reticulations, as well as by numerous diagnostic nucleotide positions in the mitochondrial 16S rRNA gene. Description of the holotype. Adult female in good state of preservation. Tissue removed from ventral side of right thigh for DNA extraction, one transverse and two longitudinal cuts on venter for gonad examination. Head longer than wide and slightly wider than body; snout rounded in dorsal, ventral and lateral views; canthus rostralis straight, loreal region flat; nostrils much nearer to tip of snout than to eye, tympanum distinct, 47% of horizontal eye diameter; supratympanic fold poorly expressed, rather straight; vomerine odontophores very poorly expressed and almost not visible, located medially between eye and choanae on either side of head; tongue ovoid, and distinctly bifid at its tip. Arms moderately thin; relative finger length 1Variation. The three morphologically examined paratypes are similar to the holotype. See Table 1 for morphological measurements. Sexual dimorphism cannot be reliably assessed as all genotyped specimens are females. Photos from specimens morphologically assigned to this species from Marojejy and Anjanaharibe-Sud agree with the overall trend of rather small dorsal spots and poorly marked purplish-brown flank patches. Etymology. The species epithet pulcherrimus is a Latin adjective meaning “most beautiful”, referring to the appealing color pattern of the new species and alluding to the name of its sister species (G. pulcher). Natural history. All known specimens were collected from the leaf axils of Pandanus screw pines in rainforest. Calls, reproductive behavior and larval stages are unknown, but it is likely that the species reproduces in these phytotelms, as does its sister species G. pulcher. Distribution. Based on genotyped specimens, G. pulcherrimus is known from (1) Hevirina and Fotsialanana source, two nearby sites on the western slope of the Makira Massif, and (2) Bemanevika. Additional specimens morphologically consistent with this taxon are known from three additional localities in northern Madagascar: (3) Anjanaharibe-Sud, (4) Marojejy, and (5) Masoala; these localities need to be genetically verified in the future., Published as part of Vences, Miguel, Hutter, Carl R., Glaw, Frank, Rakotoarison, Andolalao, Raselimanana, Achille P. & Scherz, Mark D., 2023, A new species of Pandanus-dwelling frog from northern Madagascar related to Guibemantis pulcher, pp. 97-115 in Zootaxa 5306 (1) on pages 108-112, DOI: 10.11646/zootaxa.5306.1.4, http://zenodo.org/record/8061262

Guibemantis (Pandanusicola) pulcherrimus sp. nov. Figs. 7‒9 Holotype. ZSM 518/2009 (field number ZCMV 11217), adult female, collected by M. Vences, D.R. Vieites, F.M. Ratsoavina, R.D. Randrianiaina, E. Rajeriarison, T. Rajoafiarison and J.L. Patton on 23 June 2009 near a site locally named Hevirina, western slope of the Makira Massif, Analanjirofo Region, Madagascar (GPS coordinates not taken, but coordinates of Hevirina are 15.4490°S, 49.1119°E, 1093 m a.s.l.). Paratypes. Eight specimens. ZSM 519/2009 (ZCMV 11249) and ZSM 520/2009 (ZCMV 11264), two adult females, collected by M. Vences, D.R. Vieites, F.M. Ratsoavina, R.D. Randrianiaina, E. Rajeriarison, T. Rajoafiarison and J.L. Patton on 23–24 June 2009 at a campsite near the source of Fotsialanana river, western slope of the Makira Massif, Analanjirofo Region, Madagascar (coordinates 15.46675°S, 49.128889°E, 1067 m a.s.l.); ZSM 1754/2010 (ZCMV 12505), adult female, collected by M. Vences, D.R. Vieites, R.D. Randrianiaina, F.M. Ratsoavina, S. Rasamison, A. Rakotoarison, E. Rajeriarison and T. Rajoafiarison on 27 June 2010 at Bemanevika, campsite Atsirakala, Sofia Region, Madagascar (coordinates 14.43061°S, 48.60179°E, 1466 m a.s.l.); as well as five specimens (field numbers ZCMV 11225, ZCMV 11226, ZCMV 11228, ZCMV 11229), all collected in June 2009 by by M. Vences, D.R. Vieites, R.D. Randrianiaina, F.M. Ratsoavina, S. Rasamison, A. Rakotoarison, E. Rajeriarison and T. Rajoafiarison on the western slope the Makira Massif, not examined morphologically but included in the 16S tree (Fig. 1), integrated but still uncatalogued at the UADBA (MZBA) collection (individually identifiable via their field number tags). Diagnosis. Assigned to the subgenus Pandanusicola of the genus Guibemantis based on its small body size, phytotelm-dwelling habitats (in Pandanus plants), near-absent webbing between toes, connected lateral metatarsalia, the presence of both inner and outer metatarsal tubercles, intercalary elements between ultimate and penultimate phalanges of fingers and toes (verified by external examination), and molecular phylogeny. The new species is characterized by the unique combination of the following characters: (1) medium size with SVL 25‒29 mm in females, (2) bright green dorsal color with translucent shade, with dense fine purplish-brown spots, (3) distinct brown rostral stripe, (4) absence of white or yellow spots in the inguinal region, (5) absence of webbing on hand and only traces of web at foot, (6) vomerine teeth rudimentary. The new species is most similar to its sister species G. pulcher, which differs (despite some overlap) by fewer (12–107, mean 42; vs. 47–223, mean 103) and larger (diameter often up to 0.8 mm vs. 0.4 mm) purplish-brown dorsal spots, and a larger and denser purplish-brown band on the flanks which often is adorned centrally by silvery whitish reticulations, as well as by numerous diagnostic nucleotide positions in the mitochondrial 16S rRNA gene. Description of the holotype. Adult female in good state of preservation. Tissue removed from ventral side of right thigh for DNA extraction, one transverse and two longitudinal cuts on venter for gonad examination. Head longer than wide and slightly wider than body; snout rounded in dorsal, ventral and lateral views; canthus rostralis straight, loreal region flat; nostrils much nearer to tip of snout than to eye, tympanum distinct, 47% of horizontal eye diameter; supratympanic fold poorly expressed, rather straight; vomerine odontophores very poorly expressed and almost not visible, located medially between eye and choanae on either side of head; tongue ovoid, and distinctly bifid at its tip. Arms moderately thin; relative finger length 1 After 14 years in preservative, dorsally beige with many small spots; these spots are light brown with a weakly recognizable reddish/purple shade. From the snout tip over the nostrils to the eye, and then again from the eye over the tympanum to an area above the forelimb insertions, a distinctly delimited dark brown rostral stripe is present. After the forelimb insertion, this stripe becomes broader, lighter and less distinctly delimited, and forms a brown band on the anterior flanks, dissolving into dense brown spotting on the posterior flanks. The ventral side is uniformly beige without any pattern. In life (Fig. 7A) the dorsal side was green with a translucent shade, and the spots and bands were purplish-brown. Variation. The three morphologically examined paratypes are similar to the holotype. See Table 1 for morphological measurements. Sexual dimorphism cannot be reliably assessed as all genotyped specimens are females. Photos from specimens morphologically assigned to this species from Marojejy and Anjanaharibe-Sud agree with the overall trend of rather small dorsal spots and poorly marked purplish-brown flank patches. Etymology. The species epithet pulcherrimus is a Latin adjective meaning “most beautiful”, referring to the appealing color pattern of the new species and alluding to the name of its sister species (G. pulcher). Natural history. All known specimens were collected from the leaf axils of Pandanus screw pines in rainforest. Calls, reproductive behavior and larval stages are unknown, but it is likely that the species reproduces in these phytotelms, as does its sister species G. pulcher. Distribution. Based on genotyped specimens, G. pulcherrimus is known from (1) Hevirina and Fotsialanana source, two nearby sites on the western slope of the Makira Massif, and (2) Bemanevika. Additional specimens morphologically consistent with this taxon are known from three additional localities in northern Madagascar: (3) Anjanaharibe-Sud, (4) Marojejy, and (5) Masoala; these localities need to be genetically verified in the future.

Vences, Miguel, Hutter, Carl R., Glaw, Frank, Rakotoarison, Andolalao, Raselimanana, Achille P., Scherz, Mark D. (2023): A new species of Pandanus-dwelling frog from northern Madagascar related to Guibemantis pulcher. Zootaxa 5306 (1): 97-115, DOI: 10.11646/zootaxa.5306.1.4, URL: http://dx.doi.org/10.11646/zootaxa.5306.1.4

Health-care systems use AI-driven data analytics to target high-cost patients for early interventions. Many ethicists see these programs as enacting a preferential option for the poor. Ethnographic studies, however, find that their data analytic framework emphasizes efficiency, cost containment, and constant evaluation of patients. Ongoing evaluation and surveillance can undermine other goals of Catholic health care like personal encounter and accompaniment. While targeted care programs can be implemented well, the use of AI in data analytics to serve the poor creates dangers of depersonalization.

In the last seven years, over twenty thousand people have died trying to reach Europe by crossing the Mediterranean Sea. Rescue missions by private actors and NGOs have increased because both national measures and measures by the EU’s border control agency, Frontex, are often deemed insufficient. However, such independent rescue missions face increasing persecution from national governments, Italy being one example. This raises the question of how potential migrants and dissenting citizens should act towards the EU border regime. In contrast to the literature, which mainly addresses migration on the basis of justice requirements, this article focuses on the legitimate authority of the EU’s border regime. Focusing on the legitimacy criteria for states’ claims to regulate migration opens a fruitful normative perspective, given the pervasive disagreement over the content of justice in migration. What reasons for compliance and non-interference does legitimacy supply for potential immigrants and dissenting citizens? And what legitimacy standard may be appropriate for the power that individual states claim over potential immigrants? We argue that, even assuming a minimal legitimacy standard for the state-migrant relationship, the structure of the EU’s border regime exhibits unique features, which cause it to stand in tension with such a standard. By coordinating its Member States’ border regimes, especially through Frontex, the EU claims and exercises power over potential immigrants. However, the asymmetrical delegation of state powers to the EU means that the power involved in regulating European borders is, in core respects, unaccountable. This unaccountability, we argue, is significant for the legitimacy of the EU’s border regime. This article sheds new light on the morality of unauthorised rescue missions by assessing the permissibility of resistance to the EU’s border regime.

Background Various bacterial and viral assemblages composing Cystic Fibrosis (CF) lung microbiota contribute to long-term lung function decline over time. Yet, the impact of individual microorganisms on pulmonary functions remains uncertain in children with CF. Methods As part of the 'Mucoviscidosis, respiratory VIruses, intracellular Bacteria and fastidious organisms'' project, children with CF were longitudinally followed in a Swiss multicentric study. Respiratory samples included mainly throat swabs and sputa samples for bacterial culture and 16S rRNA metagenomics and nasopharyngeal swabs for respiratory virus detection by molecular assays. Percentage of predicted Forced Expiratory Volume in one second (FEV1%) and Lung Clearance Index (LCI) were recorded. Results Sixty-one children, of whom 20 (32.8%) presented with at least one pulmonary exacerbation, were included. Almost half of the 363 nasopharyngeal swabs tested by RT-PCR were positive for a respiratory virus, mainly rhinovirus (26.5%). From linear mixed-effects regression models, P. aeruginosa (-11.35, 95%CI [-17.90; -4.80], p = 0.001) was significantly associated with a decreased FEV1%, whereas rhinovirus was associated with a significantly higher FEV1% (+4.24 95%CI [1.67; 6.81], p = 0.001). Compared to conventional culture, 16S rRNA metagenomics showed a sensitivity and specificity of 80.0% and 85.4%, respectively for detection of typical CF pathogens. However, metagenomics detected a bacteria almost twice more often than culture. Conclusions As expected, P. aeruginosa impacted negatively on FEV1% while rhinovirus was surprisingly associated with better FEV1%. Culture-free assays identifies significantly more pathogens than standard culture, with disputable clinical correlation.

PURPOSE Tumor-intrinsic features may render large B-cell lymphoma (LBCL) insensitive to CD19-directed chimeric antigen receptor T cells (CAR-T). We hypothesized that TP53 genomic alterations are detrimental to response outcomes in LBCL treated with CD19-CAR-T. MATERIALS AND METHODS Patients with LBCL treated with CD19-CAR-T were included. Targeted next-generation sequencing was performed on pre–CAR-T tumor samples in a subset of patients. Response and survival rates by histologic, cytogenetic, and molecular features were assessed. Within a cohort of newly diagnosed LBCL with genomic and transcriptomic profiling, we studied interactions between cellular pathways and TP53 status. RESULTS We included 153 adults with relapsed or refractory LBCL treated with CD19-CAR-T (axicabtagene ciloleucel [50%], tisagenlecleucel [32%], and lisocabtagene maraleucel [18%]). Outcomes echoed pivotal trials: complete response (CR) rate 54%, median overall survival (OS) 21.1 months (95% CI, 14.8 to not reached), and progression-free survival 6 months (3.4 to 9.7). Histologic and cytogenetic LBCL features were not predictive of CR. In a subset of 82 patients with next-generation sequencing profiling, CR and OS rates were comparable with the unsequenced cohort. TP53 alterations (mutations and/or copy number alterations) were common (37%) and associated with inferior CR and OS rates in univariable and multivariable regression models; the 1-year OS in TP53-altered LBCL was 44% (95% CI, 29 to 67) versus 76% (65 to 89) in wild-type ( P = .012). Transcriptomic profiling from a separate cohort of patients with newly diagnosed lymphoma (n = 562) demonstrated that TP53 alterations are associated with dysregulation of pathways related to CAR-T-cell cytotoxicity, including interferon and death receptor signaling pathway and reduced CD8 T-cell tumor infiltration. CONCLUSION TP53 is a potent tumor-intrinsic biomarker that can inform risk stratification and clinical trial design in patients with LBCL treated with CD19-CAR-T. The role of TP53 should be further validated in independent cohorts.

The small arboreal frog Guibemantis liber (Anura: Mantellidae) has served as an example for the existence of deep conspecific lineages that differ by a substantial amount in mitochondrial DNA but are similar in morphology and bioacoustics and thus are assigned to the same nominal species. During fieldwork in northern Madagascar, we identified additional such lineages and surprisingly, observed close syntopy of two of these at various sites. In-depth study based on DNA sequences of the mitochondrial cytochrome b gene from 338 specimens of G. liber sensu lato from across its range, sequences of four nuclear-encoded markers for 154‒257 of these specimens, a phylogenomic dataset obtained by the FrogCap target capture approach, and additional mitochondrial genes for representatives of most mitochondrial lineages, as well as bioacoustic and morphological comparisons, revealed concordant differentiation among several lineages of the G. liber complex. We identify nine lineages differing by 5.3‒15.5% in cytochrome b and 2.4‒10.1% in the 16S rRNA gene, and find that several of these lack or have only limited allele sharing in the nuclear-encoded genes. Based on sympatric or parapatric occurrence without genetic admixture, combined with differences in bioacoustic and morphological characters, we scientifically name three lineages from northern Madagascar as new species: G. razoky sp. nov., G. razandry sp. nov., and G. fotsitenda sp. nov. Of these new species, G. razoky sp. nov. and G. razandry sp. nov. show widespread syntopy across northern Madagascar and differ in body size and advertisement calls. Guibemantis fotsitenda sp. nov. is sister to G. razandry sp. nov., but appears to occur at lower elevations, including in close geographic proximity on the Marojejy Massif. We also detected subtle differences in advertisement calls among various other mitochondrial lineages distributed in the Northern Central East and Southern Central East of Madagascar, but the status and nomenclatural identity of these lineages require further morphological and bioacoustic study of reliably genotyped individuals, and assignment of the three available names in the complex: Rhacophorus liber Peracca, 1893, Gephyromantis albogularis Guibé, 1947, and Gephyromantis variabilis Millot and Guibé, 1951. We discuss the identity and type material of these three nomina, designate a lectotype for Gephyromantis variabilis from Itremo, and flag the collection of new material from their type localities, Andrangoloaka and Itremo, as paramount for a comprehensive revision of the G. liber complex., Field expeditions were supported, among other funders, by the Volkswagen Foundation, Deutsche Forschungsgemeinschaft, and Deutscher Akademischer Austauschdienst. A research stay of TK at TU Braunschweig was supported by a grant of the Alexander-Koenig-Gesellschaft. MP was supported by the Alexander von Humboldt Foundation. Fieldwork for CRH was supported by Global Wildlife Conservation Grant 5019-0096.

Purpose The greenhouse gas (GHG) emissions caused by the construction industry account for an enormous share of total global CO2 emissions. The numerous construction activities therefore continue to reduce the remaining carbon budget. One lever for the reduction of these GHG emissions lies in the procurement process of buildings. For this reason, a process model was developed that takes embodied and operational emissions into account in the tendering and awarding phase of buildings. Methods To validate the developed theoretical framework, environmental life cycle costing (eLCC) was conducted on a single-family house case study, taking into account external cost caused by GHG emissions. Various shadow prices were defined for the calculation of external cost to identify changes in award decisions. We further investigated a results-based climate finance (RBCF) instrument, i.e., the GHG emission bonus/malus, to demonstrate an approach for calculating Paris-compatible cost (PCC) scenarios. Results We show that an award decision based on life cycle costing (LCC) leads to a 12% reduction in GHG emissions. A further reduction in GHG emissions can be achieved by awarding contracts based on eLCC. However, the required shadow prices within the eLCC awards to influence the award decision are quite high. With the development of the LCA-based bonus/malus system, PCC scenarios can be determined at sufficient shadow prices, and further GHG emission reductions can be achieved. Conclusions Since the implementation of LCA and LCC in the tendering and awarding process is currently not mandatory, in this context, the next step towards Paris-compatible buildings must first be taken by the awarding authorities as well as the policy-makers. However, the application of the LCA-based bonus/malus system and thus the awarding of contracts according to PCC scenarios show the enormous GHG emissions reduction potential and thus represent an innovative and sustainable framework for an adapted procurement process.

supplemental materials and methods and supplemental figures

Metastasis is the main cause of cancer-related mortality. Despite intense efforts to understand the mechanisms underlying the metastatic process, treatment of metastatic cancer is still challenging. Here we describe a chemotherapy-induced, host-mediated mechanism that promotes remodeling of the extracellular matrix (ECM), ultimately facilitating cancer cell seeding and metastasis. Paclitaxel (PTX) chemotherapy enhanced rapid ECM remodeling and mechanostructural changes in the lungs of tumor-free mice, and the protein expression and activity of the ECM remodeling enzyme lysyl oxidase (LOX) increased in response to PTX. A chimeric mouse model harboring genetic LOX depletion revealed chemotherapy-induced ECM remodeling was mediated by CD8+ T cells expressing LOX. Consistently, adoptive transfer of CD8+ T cells, but not CD4+ T cells or B cells, from PTX-treated mice to naïve immunodeprived mice induced pulmonary ECM remodeling. Lastly, in a clinically relevant metastatic breast carcinoma model, LOX inhibition counteracted the metastasis-promoting, ECM-related effects of PTX. This study highlights the role of immune cells in regulating ECM and metastasis following chemotherapy, suggesting that inhibiting chemotherapy-induced ECM remodeling represents a potential therapeutic strategy for metastatic cancer.Significance:Chemotherapy induces prometastatic pulmonary ECM remodeling by upregulating LOX in T cells, which can be targeted with LOX inhibitors to suppress metastasis.See related commentary by Kolonin and Woodward, p. 197

Purpose: DOTA-AR, a bombesin-antagonist peptide, has potential clinical application for targeted imaging and therapy in gastrin-releasing peptide receptor (GRPr)–positive malignancies when conjugated with a radioisotope such as 90Y. This therapeutic potential is limited by the fast washout of the conjugates from the target tumors. WST-11 (Weizmann STeba-11 drug; a negatively charged water-soluble palladium-bacteriochlorophyll derivative, Tookad Soluble) vascular targeted photodynamic therapy (VTP) is a local ablation approach recently approved for use in early-stage prostate cancer. It generates reactive oxygen/nitrogen species within tumor blood vessels, resulting in their instantaneous destruction followed by rapid tumor necrosis. We hypothesize that the instantaneous arrest of tumor vasculature may provide a means to trap radiopharmaceuticals within the tumor, thereby improving the efficacy of targeted radiotherapy.Experimental Design: GRPr-positive prostate cancer xenografts (PC-3 and VCaP) were treated with 90Y-DOTA-AR with or without VTP. The uptake of radioisotopes was monitored by Cherenkov luminescence imaging (CLI). The therapeutic efficacy of the combined VTP and 90Y-DOTA-AR in PC-3 xenografts was assessed.Results: CLI of 90Y-DOTA-AR demonstrated longer retention of radiotracer within the VTP-treated PC-3 xenografts compared with the non–VTP-treated ones (P < 0.05) at all time points (24–144 hours) after 90Y-DOTA-AR injection. A similar pattern of retention was observed in VCaP xenografts. When 90Y-DOTA-AR administration was combined with VTP, tumor growth delay was significantly longer than for the control or the monotherapy groups.Conclusions: Tumor vascular arrest by VTP improves 90Y-DOTA-AR retention in the tumor microenvironment thereby enhancing therapeutic efficacy. Clin Cancer Res; 23(13); 3343–51. ©2017 AACR.

Patient data and univariate analysis of gastric cancer patient cohorts

RNA-seq data of PDPN+ cancer-associated fibroblasts (CAFs) from triple-transgenic gastric cancer mice

HALLMARK gene set

RNA-seq results of stroma and cancer from gastric cancer patients

ADT reduces tumor proliferation measured by Ki67

C2_CP gene set

Purpose: WST11 vascular targeted photodynamic therapy (VTP) is a local ablation approach relying upon rapid, free radical-mediated destruction of tumor vasculature. A phase III trial showed that VTP significantly reduced disease progression when compared with active surveillance in patients with low-risk prostate cancer. The aim of this study was to identify a druggable pathway that could be combined with VTP to improve its efficacy and applicability to higher risk prostate cancer tumors.Experimental Design: Transcriptome analysis of VTP-treated tumors (LNCaP-AR xenografts) was used to identify a candidate pathway for combination therapy. The efficacy of the combination therapy was assessed in mice bearing LNCaP-AR or VCaP tumors.Results: Gene set enrichment analysis identifies the enrichment of androgen-responsive gene sets within hours after VTP treatment, suggesting that the androgen receptor (AR) may be a viable target in combination with VTP. We tested this hypothesis in mice bearing LNCaP-AR xenograft tumors by using androgen deprivation therapy (ADT), degarelix, in combination with VTP. Compared with either ADT or VTP alone, a single dose of degarelix in concert with VTP significantly inhibited tumor growth. A sharp decline in serum prostate-specific antigen (PSA) confirmed AR inhibition in this group. Tumors treated by VTP and degarelix displayed intense terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling staining 7 days after treatment, supporting an increased apoptotic frequency underlying the effect on tumor inhibition.Conclusions: Improvement of local tumor control following androgen deprivation combined with VTP provides the rationale and preliminary protocol parameters for clinical trials in patients presented with locally advanced prostate cancer. Clin Cancer Res; 24(10); 2408–16. ©2018 AACR.

Kidneys and pancreas from mice bearing PC-3 tumors in 4 different treatment groups (n= 5 per each treatment group) were collected at 41 days post treatment and subjected for H&E staining to determine potential tissue toxicity caused by 90Y-DOTA-AR/VTP combination therapy. No significant lesions and no evidence of treatment induced lesions were observed in these organs (representative H&E pictures shown).

Cancer cells recruit and rewire normal fibroblasts in their microenvironment to become protumorigenic cancer-associated fibroblasts (CAF). These CAFs are genomically stable, yet their transcriptional programs are distinct from those of their normal counterparts. Transcriptional regulation plays a major role in this reprogramming, but the extent to which epigenetic modifications of DNA also contribute to the rewiring of CAF transcription is not clear. Here we address this question by dissecting the epigenetic landscape of breast CAFs. Applying tagmentation-based whole-genome bisulfite sequencing in a mouse model of breast cancer, we found that fibroblasts undergo massive DNA methylation changes as they transition into CAFs. Transcriptional and epigenetic analyses revealed RUNX1 as a potential mediator of this process and identified a RUNX1-dependent stromal gene signature. Coculture and mouse models showed that both RUNX1 and its stromal signature are induced as normal fibroblasts transition into CAFs. In breast cancer patients, RUNX1 was upregulated in CAFs, and expression of the RUNX1 signature was associated with poor disease outcome, highlighting the relevance of these findings to human disease. This work presents a comprehensive genome-wide map of DNA methylation in CAFs and reveals a previously unknown facet of the dynamic plasticity of the stroma.Significance:The first genome-wide map of DNA methylation in breast cancer–associated fibroblasts unravels a previously unknown facet of the dynamic plasticity of the stroma, with far-reaching therapeutic implications.

Gene set enrichment analysis (GSEA) using MSigDB (Hallmark gene sets) on the full stromal patient RNA-seq data

CD31 vessel staining of tumors treated with VTP or VTP/ADT combination

Distances of patient samples in PCA analysis

Pathway analysis of stroma from gastric cancer patients

Supplementary Table from Global DNA Methylation Analysis of Cancer-Associated Fibroblasts Reveals Extensive Epigenetic Rewiring Linked with RUNX1 Upregulation in Breast Cancer Stroma

Pathway analysis of PDPN+cancer-associated fibroblasts (CAFs) from triple-transgenic gastric cancer mice

Clinical characteristics of the gastric cancer patient cohort

This file contains Supplementary Materials and Methods, Supplementary Figure Legends, and 6 Supplementary Figures with their legends

Univariate and multivariate analysis of the gastric cancer patient cohort

Supplementary Data from Global DNA Methylation Analysis of Cancer-Associated Fibroblasts Reveals Extensive Epigenetic Rewiring Linked with RUNX1 Upregulation in Breast Cancer Stroma

Summary of Kaplan-Meier curves from the TCGA cohort