Ovarian carcinomas with mutations in the tumour suppressor BRCA2 are particularly sensitive to platinum compounds. However, such carcinomas ultimately develop cisplatin resistance. The mechanism of that resistance is largely unknown. Here we show that acquired resistance to cisplatin can be mediated by secondary intragenic mutations in BRCA2 that restore the wild-type BRCA2 reading frame. First, in a cisplatin-resistant BRCA2-mutated breast-cancer cell line, HCC1428, a secondary genetic change in BRCA2 rescued BRCA2 function. Second, cisplatin selection of a BRCA2-mutated pancreatic cancer cell line, Capan-1 (refs 3, 4), led to five different secondary mutations that restored the wild-type BRCA2 reading frame. All clones with secondary mutations were resistant both to cisplatin and to a poly(ADP-ribose) polymerase (PARP) inhibitor (AG14361). Finally, we evaluated recurrent cancers from patients whose primary BRCA2-mutated ovarian carcinomas were treated with cisplatin. The recurrent tumour that acquired cisplatin resistance had undergone reversion of its BRCA2 mutation. Our results suggest that secondary mutations that restore the wild-type BRCA2 reading frame may be a major clinical mediator of acquired resistance to platinum-based chemotherapy.

The mutational spectra of BRCA1 and BRCA2 include many high-penetrance, individually rare genomic rearrangements. Among patients with breast cancer and severe family histories of cancer who test negative (wild type) for BRCA1 and BRCA2, approximately 12% can be expected to carry a large genomic deletion or duplication in one of these genes, and approximately 5% can be expected to carry a mutation in CHEK2 or TP53. Effective methods for identifying these mutations should be made available to women at high risk.

Large liquid helium clusters (He L , n+10) produced in a supersonic jet are doped with alkali atoms (Li, Na, K) and characterized by means of laser induced fluorescence. Each cluster contains, on average, less than one dopant atom. Both excitation and emission spectra have been recorded. The observed excitation spectra are analyzed, calculating the transitions within an approach based on the hypothesis that the chromophores are trapped in a dimple on the cluster's surface as predicted by the theoretical calculations of Ancilotto et al. [9]. The results of the model calculations are in good qualitative agreement with the experimental findings. In spite of the very weak binding energy (a few cm\), some of the excited atoms remain bound to the surface, provided the excitation occurs at frequencies not too far from the alkali's gas phase absorptions. These bound-bound excitations produce very broad, red shifted emission spectra. At other, blue shifted frequencies, the excited atoms desorb from the cluster's surface, giving rise to unshifted, free atom, emission spectra. The heavier alkali metals (Na, K) show, compared to the calculations, an additional broadening which is attributed to surface excitations on the helium droplet.

Acquired platinum resistance is a serious problem in the treatment of ovarian carcinomas. However, the mechanism of the drug resistance has not been elucidated. Here, we show functional significance of restoration of BRCA2 protein by secondary BRCA2 mutations in acquired drug resistance of BRCA2-mutated ovarian carcinoma. Three ovarian cancer cell lines (PEO1, PEO4, and PEO6) were derived from a BRCA2 mutation [5193C>G (Y1655X)] carrier with ovarian carcinoma with acquired cisplatin resistance and a secondary BRCA2 mutation [5193C>T (Y1655Y)] that canceled the inherited mutation. PEO1 was BRCA2 deficient and sensitive to cisplatin and a poly(ADP-ribose) polymerase inhibitor, AG14361, whereas PEO4 was resistant. PEO4 and PEO6, derived from ascites at the time of relapse with cisplatin resistance, had the secondary mutation and were BRCA2 proficient. In vitro cisplatin/AG14361 selection of PEO1 led to restoration of BRCA2 due to another secondary BRCA2 mutation. BRCA2 depletion sensitized BRCA2-restored PEO1 clones and PEO4 to cisplatin/AG14361. Thus, restoration of BRCA2 due to secondary BRCA2 mutation is involved in acquired drug resistance of BRCA2-mutated ovarian carcinoma. [Cancer Res 2009;69(16):6381-6]

Helium cluster isolation spectroscopy is a recently developed spectroscopic method that involves the formation of a beam of large helium clusters (104 atoms per cluster), the capture by the clusters of the atoms or molecules of interest in a low-pressure pick-up cell, and the spectroscopic study of the isolated species. Here we exploit the unique feature of this method of allowing the selective preparation of high-spin molecular species (e.g., triplet dimers) over their low-spin (singlet) counterparts to measure the spectra of several alkali dimers in their triplet manifold. By probing via laser-induced fluorescence their lowest triplet-to-triplet transitions, Li2, Na2, K2, and NaK are found to reside on the surface of the helium clusters. Since the spectroscopic shifts induced by the helium cluster are minimal, vibrational analysis of the electronic transitions produces transition frequencies that can be compared to previous ab initio and experimental values. Both bound−bound and bound−free transitions have been observed. Emission spectra reveal the presence of vibrational relaxation and nonadiabatic intersystem crossings of the excited dimers that result from the proximity of the helium cluster surface. Through this study we improve our understanding of triplet alkali dimer potential energy curves, we test an efficient analytical model to represent them, and we provide input information for the study of nonadditive effects present in quartet (spin-polarized) alkali trimers which can be formed using the same method.

Nanometer-sized helium droplets, each containing about 10(4) helium atoms, were used as an inert substrate on which to form previously unobserved, spin-3/2 (quartet state) alkali trimers. Dispersed fluorescence measurements reveal that, upon electronic excitation, the quartet trimers undergo intersystem crossing to the doublet manifold, followed by dissociation of the doublet trimer into an atom and a covalently bound singlet dimer. As shown by this work, aggregates of spin-polarized alkali metals represent ideal species for the optical study of fundamental chemical dynamics processes including nonadiabatic spin conversion, change of bonding nature, and unimolecular dissociation.

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In this paper we describe an application of reversed time-correlated single photon counting to the time-resolved spectroscopy of impurity atoms and molecules bound to large quantum clusters. The photo-induced dynamics of Na atoms on the surface of He and clusters have been studied by follow-H 2 ing the time dependence of their emission at selected excitation and emission wavelengths. Collection of atomic (16 980 ^145 cm~1) Ñuorescence arising from Na atoms excited on the cluster surface and immediately desorbed from it yields a Ðnite (ca. 70 ps) rise time and a decay time of 16.3 ^0.1 ns, equal to the known lifetime of the 3P ] 3S transition of atomic Na. The frequency distribution of the red emission due to atoms that do not leave the cluster immediately after excitation is shown to be due to a desorbed Na*ÈHe exciplex by obtaining quantitative agreement with predictions derived from available ab initio NaÈHe potentials. Formation of this excimer can occur along either the or excited state surface. 2% 1@2 2% 3@2 " Slow Ï (700 ps) and " fast Ï (ca. 70 ps) components of the rise time of the red emission (15 800 ^125 cm~1) are assigned, respectively, to the two formation channels. Introducing spinÈorbit coupling e †ects into the long range ab initio pair potential for an isolated Na*ÈHe generates a small barrier on the potential curve, which is linked to the observed slow exciplex forma-2% 1@2 tion time.