Genetic reversal of the globin switch concurrently modulates both fetal and sickle hemoglobin and reduces red cell sickling.

We previously reported initial clinical results of post-transcriptional gene silencing of BCL11A expression (NCT03282656) reversing the fetal to adult hemoglobin switch. A goal of this approach is to increase fetal hemoglobin (HbF) expression while coordinately reducing sickle hemoglobin (HbS) expression. The resulting combinatorial effect should prove effective in inhibiting HbS polymerization at lower physiologic oxygen values thereby mitigating disease complications. Here we report results of exploratory single-cell analysis of patients in which BCL11A is targeted molecularly and compare results with cells of patients treated with hydroxyurea (HU), the current standard of care. We use single-cell assays to assess HbF, HbS, oxygen saturation, and hemoglobin polymer content in RBCs for nine gene therapy trial subjects (BCLshmiR, median HbF% = 27.9) and compare them to 10 HU-treated subjects demonstrating high and comparable levels of HbF (HU High Responders, median HbF% = 27.0). All BCL11A patients achieved the primary endpoint for NCT03282656, which was defined by an absolute neutrophil count greater than or equal to 0.5 × 109 cells/L for three consecutive days, achieved within 7 weeks following infusion. Flow cytometric assessment of single-RBC HbF and HbS shows fewer RBCs with high HbS% that would be most susceptible to sickling in BCLshmiR vs. HU High Responders: median 42% of RBCs with HbS%>70% in BCLshmiR vs. 61% in HU High Responders (p = 0.004). BCLshmiR subjects also demonstrate more RBCs resistant to HbS polymerization at lower physiologic oxygen tension: median 32% vs. 25% in HU High Responders (p = 0.006). Gene therapy-induced BCL11A down-regulation reverses the fetal-to-adult hemoglobin switch and induces RBCs with higher HbF%, lower HbS%, and greater resistance to deoxygenation-induced polymerization in clinical trial subjects compared with a cohort of highly responsive hydroxyurea-treated subjects.

Long-term outcomes of lentiviral gene therapy for the ß-hemoglobinopathies: the HGB-205 trial.

Sickle cell disease (SCD) and transfusion-dependent ß-thalassemia (TDT) are the most prevalent monogenic disorders worldwide. Trial HGB-205 ( NCT02151526 ) aimed at evaluating gene therapy by autologous CD34+ cells transduced ex vivo with lentiviral vector BB305 that encodes the anti-sickling ßA-T87Q-globin expressed in the erythroid lineage. HGB-205 is a phase 1/2, open-label, single-arm, non-randomized interventional study of 2-year duration at a single center, followed by observation in long-term follow-up studies LTF-303 ( NCT02633943 ) and LTF-307 ( NCT04628585 ) for TDT and SCD, respectively. Inclusion and exclusion criteria were similar to those for allogeneic transplantation but restricted to patients lacking geno-identical, histocompatible donors. Four patients with TDT and three patients with SCD, ages 13-21 years, were treated after busulfan myeloablation 4.6-7.9 years ago, with a median follow-up of 4.5 years. Key primary endpoints included mortality, engraftment, replication-competent lentivirus and clonal dominance. No adverse events related to the drug product were observed. Clinical remission and remediation of biological hallmarks of the disease have been sustained in two of the three patients with SCD, and frequency of transfusions was reduced in the third. The patients with TDT are all transfusion free with improvement of dyserythropoiesis and iron overload.

Correcting photodetector nonlinearity in dual-comb interferometry.

Photodetector nonlinearity, the main limiting factor in terms of optical power in the detection chain, is corrected to improve the signal-to-noise ratio of a short-time measurement in dual-comb spectroscopy. An iterative correction algorithm minimizing out-of-band spectral artifacts based on nonlinearity correction methods used in classical Fourier-transform spectrometers is presented. The exactitude of the nonlinearity correction is validated using a low power linear measurement. Spectroscopic lines of H12CN are provided and the increase in absorption depth of 24% caused by the saturation of the detector is corrected yielding residuals limited by the measurement noise.

Orthogonalizing the control of frequency combs for optical clockworks.

Frequency combs play a crucial supporting role for optical clocks by allowing coherent frequency division of their output signals into the electronic domain. This task requires stabilization of the comb's offset frequency and of an optical comb mode to the clock laser. However, the two actuators used to control these quantities often influence both degrees of freedom simultaneously. This non-orthogonality leads to artificial limits to the control bandwidth and unwanted noise in the comb. Here, we orthogonalize the two feedback loops with a linear combination of the measured signals in a field-programmable gate array. We demonstrate this idea using a fiber frequency comb stabilized to a clock laser at 259 THz, half the frequency of the 1S0→3P0 Yb transition. The decrease in coupling between the loops reduces the comb's optical phase noise by 20 dB. This approach could improve the performance of any comb stabilized to any optical frequency standard.

Hertz-level frequency comparisons between diverse color lasers without a frequency comb.

We present a simple yet powerful technique to measure and stabilize the relative frequency noise between two lasers emitting at vastly different wavelengths. The noise of each laser is extracted simultaneously by a frequency discriminator built around an unstabilized Mach-Zehnder fiber interferometer. Our protocol ensures that the instability of the interferometer is canceled and yields a direct measure of the relative noise between the lasers. As a demonstration, we measure the noise of a 895 nm diode laser against a reference laser located hundreds of nm away at 1561 nm. We also demonstrate the ability to stabilize the two lasers with a control bandwidth of 100 kHz using a Red Pitaya and reach a sensitivity of 1Hz2/Hz limited by detector noise. We independently verify the performance using a commercial frequency comb. This approach stands as a simple and cheap alternative to frequency combs to transport frequency stability across large spectral intervals or to characterize the noise of arbitrary color sources.

Individual red blood cell fetal hemoglobin quantification allows to determine protective thresholds in sickle cell disease.

Polymerization of the sickle hemoglobin (HbS) is a key determinant of sickle cell disease (SCD), an inherited blood disorder. Fetal hemoglobin (HbF) is a major modulator of the disease severity by both decreasing HbS intracellular concentration and inhibiting its polymerization. However, heterocellular distribution of HbF is common in SCD. For HbS polymerization inhibition, the hypothesis of an "HbF per red blood cell (HbF/RBC) threshold" requires accurate measurement of HbF in individual RBC. To date, HbF detection methods are limited to a qualitative measurement of RBC populations containing HbF - the F cells, which are variable. We developed an accurate method for HbF quantification in individual RBC. A linear association between mean HbF content and mean RBC fluorescence by flow cytometry, using an anti-Human-HbF antibody, was obtained from non-SCD subjects presenting homogeneous HbF distribution. This correlation was then used to measure HbF/RBC. Hydroxyurea (HU) improves SCD clinical manifestations, mainly through its ability to induce HbF synthesis. The HbF distribution was analyzed in 14 SCD patients before and during HU treatment. A significant decrease in RBC population containing less than 2 pg of HbF/RBC was observed. Therefore, we tested associations for %RBC above different HbF/RBC thresholds and showed a decrease in the pathognomonic vaso-occlusive crisis incidence from the threshold of 4 pg. This quantity was also correlated with the level of sickle RBC after in vitro deoxygenation. This new method allows the comparison of HbF/RBC distributions and could be a useful tool to characterize baseline patients HbF distribution and therapeutic response to HbF inducers.

Toward free-running operation of dual-comb fiber lasers for methane sensing.

The phase information provided by the beat note between frequency combs and two continuous-wave lasers is used to extrapolate the phase evolution of comb modes found in a spectral region obtained via nonlinear broadening. This thereafter enables using interferogram self-correction to fully retrieve the coherence of a dual-comb beat note between two independent fiber lasers. This approach allows the $ f - 2f $f-2f self-referencing of both combs, which is a significant simplification. Broadband near-infrared methane spectroscopy has been conducted to demonstrate the simplified system's preserved performance.

The internal brain drain: foreign aid, hiring practices, and international migration.

Building on empirical material gathered in Haiti, this paper advances a new and innovative understanding of the internal brain drain phenomenon-the poaching of local skilled workers by international organisations (IOs) or international non-governmental organisations (INGOs)- by conceptualising it as an equilibrium. This equilibrium is composed of two sets of tensions: (i) those between the salary conditions in the public sector and those on offer to local personnel working for IOs and INGOs; and (ii) those inherent in the dual salary scale used by IOs and INGOs for local and international staff. These two sets of tensions contribute in their specific ways to international migration, and, as such, the internal brain drain has a bearing on external brain drain dynamics. In addition, the paper addresses the difficult policy choices facing development and humanitarian organisations, since every set of policies that impacts on one side of the equilibrium is bound to affect its other side.

Single-frequency mid-infrared waveguide laser.

A guided-wave chip laser operating in a single longitudinal mode at 2860 nm is presented. The cavity was set in the Littman-Metcalf configuration to achieve single-frequency operation with a side-mode suppression ratio above 33 dB. The chip laser's 2 MHz linewidth on a 10 ms scale was found to be limited by mechanical fluctuations, but its Lorentzian contribution was estimated to be lower than 1 Hz using a heterodyne technique. This demonstration incorporates a high coherence source with the simplicity provided by the compactness of chip lasers.

Methane spectroscopy using a free-running chip-based dual-comb laser.

Absorption lines of methane in the 2ν3 band centered at 1650 nm were measured with a free-running mode-locked dual-comb laser based on a single erbium-doped glass chip. The laser's spectra were broadened up to 1670 nm using amplifiers and highly nonlinear fiber. A comb was used to interrogate the complex transmission spectrum of a methane-filled gas cell with an optical point spacing of 968 MHz and an interferogram (IGM) rate of 27 kHz to yield absorption lines of the R and Q branches. A 1.28 s sequence of IGMs was measured and phase-corrected using a self-sufficient correction algorithm seeded only by the IGMs. The associated transmission spectrum was then compared to HITRAN yielding residuals limited by photodetector nonlinearity.

Dual electro-optic frequency comb spectroscopy using pseudo-random modulation.

An approach for dual-comb spectroscopy using electro-optic (EO) phase modulation is reported. Maximum-length pseudo-random binary sequences allow for energy-efficient and flexible comb generation. Self-correction of interferograms is shown to remove relative comb drifts and improve mutual coherence, even for EO combs derived from the same laser source. Methane spectroscopy is reported over a ∼10 GHz spectral range, limited by the modulators' bandwidth. The potential of a simple EO comb instrument is demonstrated to rapidly quantify atmospheric methane emissions with ppb precision over 1 km.

An open and flexible digital phase-locked loop for optical metrology.

This paper presents an open and flexible digital phase-locked loop optimized for laser stabilization systems. It is implemented on a cheap and easily accessible FPGA-based digital electronics platform (Red Pitaya) running a customizable open-source firmware. A PC-based software interface allows controlling the platform and optimizing the loop parameters remotely. Several tools are included to allow measurement of quantities of interest smoothly and rapidly. To demonstrate the platform's capabilities, we built a fiber noise canceller over a 400 m fiber link. Noise cancellation was achieved over a 30 kHz bandwidth, a value limited mainly by the delays introduced by the actuator and by the round-trip propagation over the fiber link. We measured a total latency of 565 ns for the platform itself, limiting the theoretically achievable control bandwidth to approximately 225 kHz.

Highly coherent free-running dual-comb chip platform.

We characterize the frequency noise performance of a free-running dual-comb source based on an erbium-doped glass chip running two adjacent mode-locked waveguide lasers. This compact laser platform, contained only in a 1.2 L volume, rejects common-mode environmental noise by 20 dB thanks to the proximity of the two laser cavities. Furthermore, it displays a remarkably low mutual frequency noise floor around 10 Hz2/Hz, which is enabled by its large-mode-area waveguides and low Kerr nonlinearity. As a result, it reaches a free-running mutual coherence time of 1 s since mode-resolved dual-comb spectra are generated even on this time scale. This design greatly simplifies dual-comb interferometers by enabling mode-resolved measurements without any phase lock.

Self-corrected chip-based dual-comb spectrometer.

We present a dual-comb spectrometer based on two passively mode-locked waveguide lasers integrated in a single Er-doped ZBLAN chip. This original design yields two free-running frequency combs having a high level of mutual stability. We developed in parallel a self-correction algorithm that compensates residual relative fluctuations and yields mode-resolved spectra without the help of any reference laser or control system. Fluctuations are extracted directly from the interferograms using the concept of ambiguity function, which leads to a significant simplification of the instrument that will greatly ease its widespread adoption and commercial deployment. Comparison with a correction algorithm relying on a single-frequency laser indicates discrepancies of only 50 attoseconds on optical timings. The capacities of this instrument are finally demonstrated with the acquisition of a high-resolution molecular spectrum covering 20 nm. This new chip-based multi-laser platform is ideal for the development of high-repetition-rate, compact and fieldable comb spectrometers in the near- and mid-infrared.

Ultrafast pulse generation in a mode-locked Erbium chip waveguide laser.

We report mode-locked ~1550 nm output of transform-limited ~180 fs pulses from a large mode-area (diameter ~50 µm) guided-wave erbium fluorozirconate glass laser. The passively mode-locked oscillator generates pulses with 25 nm bandwidth at 156 MHz repetition rate and peak-power of 260 W. Scalability to higher repetition rate is demonstrated by transform-limited 410 fs pulse output at 1.3 GHz. To understand the origins of the broad spectral output, the laser cavity is simulated by using a numerical solution to the Ginzburg-Landau equation. This paper reports the widest bandwidth and shortest pulses achieved from an ultra-fast laser inscribed waveguide laser.

Dual-comb spectroscopy with a phase-modulated probe comb for sub-MHz spectral sampling.

We present a straightforward and efficient method to reduce the mode spacing of a frequency comb based on binary pseudo-random phase modulation of its pulse train. As a proof of concept, we use such a densified comb to perform dual-comb spectroscopy of a long-delay Mach-Zehnder interferometer and a high-quality-factor microresonator with sub-MHz spectral sampling. Since this approach is based on binary phase modulation, it combines all the advantages of other densification techniques: simplicity, single-step implementation, and conservation of the initial comb's power.

Complex direct comb spectroscopy with a virtually imaged phased array.

We demonstrate a simple interferometric technique to directly measure the complex optical transmittance over a large spectral range using a frequency-comb spectrometer based on a virtually imaged phased array. A Michelson interferometer encodes the phase deviations induced by a sample contained in one of its arms into an interferogram image. When combined with an additional image taken from each arm separately, along with a frequency-calibration image, this allows full reconstruction of the sample's optical transfer function. We demonstrate the technique with a vapor cell containing H13C14N, producing transmittance and phase spectra spanning 2.9 THz (∼23 nm) with ∼1 GHz resolution.

Self-heterodyne interference spectroscopy using a comb generated by pseudo-random modulation.

We present an original instrument designed to accomplish high-speed spectroscopy of individual optical lines based on a frequency comb generated by pseudo-random phase modulation of a continuous-wave (CW) laser. This approach delivers efficient usage of the laser power as well as independent control over the spectral point spacing, bandwidth and central wavelength of the comb. The comb is mixed with a local oscillator generated from the same CW laser frequency-shifted by an acousto-optic modulator, enabling a self-heterodyne detection scheme. The current configuration offers a calibrated spectrum every 1.12 µs. We demonstrate the capabilities of the spectrometer by producing averaged, as well as time-resolved, spectra of the D1 transition of cesium with a 9.8-MHz point spacing, a 50-kHz resolution and a span of more than 3 GHz. The spectra obtained after 1 ms of averaging are fitted with complex Voigt profiles that return parameters in good agreement with expected values.