Acute Myeloid Leukemia Case after Gene Therapy for Sickle Cell Disease.

Gene therapy with LentiGlobin for sickle cell disease (bb1111, lovotibeglogene autotemcel) consists of autologous transplantation of a patient's hematopoietic stem cells transduced with the BB305 lentiviral vector that encodes the ßA-T87Q-globin gene. Acute myeloid leukemia developed in a woman approximately 5.5 years after she had received LentiGlobin for sickle cell disease as part of the initial cohort (Group A) of the HGB-206 study. An analysis of peripheral-blood samples revealed that blast cells contained a BB305 lentiviral vector insertion site. The results of an investigation of causality indicated that the leukemia was unlikely to be related to vector insertion, given the location of the insertion site, the very low transgene expression in blast cells, and the lack of an effect on expression of surrounding genes. Several somatic mutations predisposing to acute myeloid leukemia were present after diagnosis, which suggests that patients with sickle cell disease are at increased risk for hematologic malignant conditions after transplantation, most likely because of a combination of risks associated with underlying sickle cell disease, transplantation procedure, and inadequate disease control after treatment. (Funded by Bluebird Bio.).

Induction of hemangiosarcoma in mice after chronic treatment with S1P-modulator siponimod and its lack of relevance to rat and human.

A high incidence of hemangiosarcoma (HSA) was observed in mice treated for 2 years with siponimod, a sphingosine-1-phosphate receptor 1 (S1P1) functional antagonist, while no such tumors were observed in rats under the same treatment conditions. In 3-month rat (90 mg/kg/day) and 9-month mouse (25 and 75 mg/kg/day) in vivo mechanistic studies, vascular endothelial cell (VEC) activation was observed in both species, but VEC proliferation and persistent increases in circulating placental growth factor 2 (PLGF2) were only seen in the mouse. In mice, these effects were sustained over the 9-month study duration, while in rats increased mitotic gene expression was present at day 3 only and PLGF2 was induced only during the first week of treatment. In the mouse, the persistent VEC activation, mitosis induction, and PLGF2 stimulation likely led to sustained neo-angiogenesis which over life-long treatment may result in HSA formation. In rats, despite sustained VEC activation, the transient mitotic and PLGF2 stimuli did not result in the formation of HSA. In vitro, the mouse and rat primary endothelial cell cultures mirrored their respective in vivo findings for cell proliferation and PLGF2 release. Human VECs, like rat cells, were unresponsive to siponimod treatment with no proliferative response and no release of PLGF2 at all tested concentrations. Hence, it is suggested that the human cells also reproduce a lack of in vivo response to siponimod. In conclusion, the molecular mechanisms leading to siponimod-induced HSA in mice are considered species specific and likely irrelevant to humans.

Niche partitioning in the coevolution of 2 distinct RNA enzymes.

Organisms that compete for limited resources within a common environment may evolve traits that allow them to exploit distinct ecological niches, thus enabling multiple species to coexist within the same habitat. The process of niche partitioning now has been captured at the molecular level, employing the method of continuous in vitro evolution. Mixed populations of 2 different "species" of RNA enzymes were made to compete for limited amounts of one or more substrates, with utilization of the substrate being necessary for amplification of the RNA. Evolution in the presence of a single substrate led to the extinction of one or the other enzyme, whereas evolution in the presence of 5 alternative substrates led to the accumulation of mutations that allowed each enzyme to exploit a different preferred resource. The evolved enzymes were capable of sustained coevolution within a common environment, exemplifying the emergence of stable ecological niche behavior in a model system. Biochemical characterization of the 2 evolved enzymes revealed marked differences in their kinetic properties and adaptive strategies. One enzyme reacted with its preferred substrate approximately 100-fold faster than the other, but the slower-reacting species produced 2- to 3-fold more progeny per reacted parent molecule. The in vitro coevolution of 2 or more species of RNA enzymes will make possible further studies in molecular ecology, including the exploration of more complex behaviors, such as predation or cooperation, under controlled laboratory conditions.

Emergence of a fast-reacting ribozyme that is capable of undergoing continuous evolution.

It is possible to evolve RNA enzymes in a continuous manner by employing a simple serial-transfer procedure. This method was previously applied only to descendants of one unusually fast-reacting RNA enzyme with RNA ligase activity. The present study establishes a second continuously evolving RNA enzyme, also with RNA ligase activity, but with a completely independent evolutionary origin. Critical to achieving the fast catalytic rate necessary for continuous evolution, development of this enzyme entailed the addition and evolutionary maturation of a 35-nucleotide accessory domain and the application of highly stringent selection pressure, with reaction times as short as 15 ms. Once established, continuous evolution was carried out for 80 successive transfers, maintaining the population against an overall dilution of 10(207)-fold. The resulting RNA enzymes exhibited approximately 10(5)-fold improvement in catalytic efficiency, compared with the starting molecules, and became dependent on a structural feature of the substrate that previously conferred no selective advantage. This adaptation was eliminated by deleting the substrate feature and then carrying out 20 additional transfers of continuous evolution, which resulted in molecules with even greater catalytic activity. Now that two distinct species of continuously evolving enzymes have been established, it is possible to conduct molecular ecology experiments in which the two are made to compete for limited resources within a common environment.