Gustavo A. Ruiz Buendía

The local and systemic symptoms that follow vaccination, collectively referred to as reactogenicity, are common, yet the mechanisms underlying individual variability remain poorly understood. Through longitudinal immune profiling of vaccinated individuals and mechanistic studies in mice, we identified key immunological determinants of reactogenicity induced by mRNA vaccines. Systemic adverse events were associated with stronger interferon and pro-inflammatory responses after the second dose of a COVID-19 mRNA vaccine, which were also correlated with the magnitude of the antigen-specific adaptive responses. This heightened inflammation occurred within 24 hours of vaccination, originated primarily from the injection site, and was characterized by enhanced recruitment and activation of myeloid cells, particularly monocytes. Two mechanisms contributed to this response: early interferon production by muscle T cells generated after the first dose and Fcγ receptor–dependent chemokine induction by vaccine antigen–specific antibodies. Consistently, serum antibody levels before vaccination correlated positively with reactogenicity. In addition to this local amplification mechanism, variability in reactogenicity was influenced by the baseline immune state, given that individuals with a preexisting interferon-stimulated gene signature in monocytes, detectable at both transcriptomic and epigenetic levels, were more prone to systemic symptoms. Together, our findings revealed molecular and cellular mechanisms driving vaccine reactogenicity, providing a framework for the design of less reactogenic vaccines.

Background New onsets of chronic urticaria (CU) have been reported after repeated immunizations, mainly with the Moderna mRNA-1273 vaccine (Spikevax). This study aims to evaluate patients with CU after COVID-19 mRNA vaccination. The contribution of SARS-Cov2 infection, atopy and IgE against the vaccine was analyzed. Methods We monitored the features of patients who developed CU after vaccination through two surveys conducted in 2022 and 2023. Fifty individuals with CU underwent blood tests, and their results were compared with individuals without a history of urticaria (N = 135). The presence of anti-vaccine IgE was tested in 185 individuals with basophil activation tests (BAT). We assessed anti-SARS-Cov2 humoral response, and the presence of IgEs against common respiratory allergens (Phadiatop) as a surrogate for atopy. Results Post-vaccination CU occurs after a median interval of 10 days and significantly more after the Spikevax booster, affecting middle-aged individuals (median 41, 66% females). In 2023, CU was still active in 53% of the cases. Inducible forms of CU, primarily dermographism, are reported in 54% (2022) and 61% (2023) of the cases. BAT positivity is not specific to CU, anti-nucleocapsid positivity, or atopy but is significantly associated with higher anti-spike neutralizing activities and younger age. Four CU patients tolerate an additional dose of mRNA vaccine with no disease exacerbation/recurrence. Conclusions The spikevax booster induces anti-vaccine IgE independently of CU, the latter being not directly associated with COVID-19 infection nor atopy. The tolerance to a new booster in 4/4 patients suggests that the Spikevax vaccine indirectly triggers CU in predisposed individuals.

Background New onsets of chronic urticaria (CU) have been reported after repeated immunizations, mainly with the Moderna mRNA-1273 vaccine (Spikevax) Objective This study aims to evaluate patients with CU after COVID-19 mRNA vaccination. The contribution of SARS-Cov2 infection, atopy and IgE against the vaccine was analyzed. Methods We monitored the features of patients who developed CU after vaccination in the Canton of Vaud through two surveys conducted in 2022 and 2023. Fifty individuals with CU underwent blood tests, and their results were compared with individuals without a history of urticaria (N=135). The presence of anti-vaccine IgE was detected with basophil activation tests (BAT). We assessed anti-SARS-Cov2 humoral response, and the presence of IgEs against common respiratory allergens (Phadiatop) as a surrogate for atopy. Results Post-vaccination CU occurred after a median interval of 10 days and significantly more after the Spikevax booster, affecting middle-aged individuals (median 41, 66% females). In 2023, CU was still active in 53% of the cases. Inducible forms of CU, primarily dermographism, were reported in 54% (2022) and 61% (2023) of the cases. BAT positivity was not specific to CU, anti-nucleocapsid positivity, or atopy but was significantly associated with higher anti-spike neutralizing activities and younger age. Four CU patients tolerated an additional dose of mRNA vaccine with no disease exacerbation/recurrence. Conclusion The Spikevax booster induced anti-vaccine IgE independently of CU, the latter being not directly associated with COVID-19 infection nor atopy. The tolerance to a new booster in 4/4 patients suggests that the Spikevax vaccine indirectly triggered CU in predisposed individuals.

Gain-of-function mutations in NOTCH1 are among the most frequent genetic alterations in T-cell acute lymphoblastic leukemia (T-ALL), highlighting the Notch signaling pathway as a promising therapeutic target for personalized medicine. Yet, a major limitation for long-term success of targeted therapy is relapse due to tumor heterogeneity or acquired resistance. Thus, we performed a genome-wide CRISPR-Cas9 screen to identify prospective resistance mechanisms to pharmacological NOTCH inhibitors and novel targeted combination therapies to efficiently combat T-ALL. Mutational loss of phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) causes resistance to Notch inhibition. PIK3R1 deficiency leads to increased PI3K/AKT signaling, which regulates cell cycle and the spliceosome machinery, both at the transcriptional and posttranslational level. Moreover, several therapeutic combinations have been identified, in which simultaneous targeting of the cyclin-dependent kinases 4 and 6 (CDK4/6) and NOTCH proved to be the most efficacious in T-ALL xenotransplantation models.

Gain-of-function mutations in NOTCH1 are among the most frequent genetic alterations in T cell acute lymphoblastic leukemia (T-ALL), making the Notch signaling pathway a promising therapeutic target for personalized medicine. Yet, a major limitation for long-term success of targeted therapy is relapse due to tumor heterogeneity or acquired resistance. Thus, we performed a genome-wide CRISPR-Cas9 screen to identify prospective resistance mechanisms to pharmacological NOTCH inhibitors and novel targeted combination therapies to efficiently combat T-ALL. Mutational loss of Phosphoinositide-3-Kinase regulatory subunit 1 (PIK3R1) causes resistance to Notch inhibition. PIK3R1 deficiency leads to increased PIK3/Akt signaling which regulates the cell cycle and spliceosome machinery, both at the transcriptional and post-translational level. Moreover, several therapeutic combinations have been identified, where simultaneous targeting of the cyclin-dependent kinases 4 and 6 (CDK4/6) and NOTCH proved to be the most efficacious in T-ALL xenotransplantation models. Key points: Mutational loss of PIK3R1 induces resistance to NOTCH1 inhibition in T-ALL Pharmacological Notch inhibition synergizes with CDK4/6 inhibitors in T-ALL

NOTCH1 is a well-established lineage specifier for T cells and among the most frequently mutated genes throughout all subclasses of T cell acute lymphoblastic leukemia (T-ALL). How oncogenic NOTCH1 signaling launches a leukemia-prone chromatin landscape during T-ALL initiation is unknown. Here we demonstrate an essential role for the high-mobility-group transcription factor Tcf1 in orchestrating chromatin accessibility and topology, allowing aberrant Notch1 signaling to convey its oncogenic function. Although essential, Tcf1 is not sufficient to initiate leukemia. The formation of a leukemia-prone epigenetic landscape at the distal Notch1-regulated Myc enhancer, which is fundamental to this disease, is Tcf1-dependent and occurs within the earliest progenitor stage even before cells adopt a T lymphocyte or leukemic fate. Moreover, we discovered a unique evolutionarily conserved Tcf1-regulated enhancer element in the distal Myc-enhancer, which is important for the transition of preleukemic cells to full-blown disease.

Expanded CAG/CTG repeats underlie 13 neurological disorders, including myotonic dystrophy type 1 (DM1) and Huntington’s disease (HD). Upon expansion, disease loci acquire heterochromatic characteristics, which may provoke changes to chromatin conformation and thereby affect both gene expression and repeat instability. Here, we tested this hypothesis by performing 4C sequencing at the DMPK and HTT loci from DM1 and HD–derived cells. We find that allele sizes ranging from 15 to 1700 repeats displayed similar chromatin interaction profiles. This was true for both loci and for alleles with different DNA methylation levels and CTCF binding. Moreover, the ectopic insertion of an expanded CAG repeat tract did not change the conformation of the surrounding chromatin. We conclude that CAG/CTG repeat expansions are not enough to alter chromatin conformation in cis. Therefore, it is unlikely that changes in chromatin interactions drive repeat instability or changes in gene expression in these disorders.

We present µLAS, a lab-on-chip system that concentrates, separates, and detects DNA fragments in a single module. µLAS speeds up DNA size analysis in minutes using femtomolar amounts of amplified DNA. Here we tested the relevance of µLAS for sizing expanded trinucleotide repeats, which cause over 20 different neurological and neuromuscular disorders. Because the length of trinucleotide repeats correlates with the severity of the diseases, it is crucial to be able to size repeat tract length accurately and efficiently. Expanded trinucleotide repeats are however genetically unstable and difficult to amplify. Thus, the amount of amplified material to work with is often limited, making its analysis labor-intensive. We report the detection of heterogeneous allele lengths in 8 samples from myotonic dystrophy type 1 and Huntington disease patients with up to 750 CAG/CTG repeats in five minutes or less. The high sensitivity of the method allowed us to minimize the number of amplification cycles and thus reduce amplification artefacts without compromising the detection of the expanded allele. These results suggest that µLAS can speed up routine molecular biology applications of repetitive sequences and may improve the molecular diagnostic of expanded repeat disorders.

The natural restoration of soils polluted by aromatic hydrocarbons such as benzene, toluene, ethylbenzene and m- and p-xylene (BTEX) may be accelerated by inoculation of specific biodegraders (bioaugmentation). Bioaugmentation mainly involves introducing bacteria that deploy their metabolic properties and adaptation potential to survive and propagate in the contaminated environment by degrading the pollutant. In order to better understand the adaptive response of cells during a transition to contaminated material, we analyzed here the genome and short-term (1 h) changes in genome-wide gene expression of the BTEX-degrading bacterium Pseudomonas veronii 1YdBTEX2 in non-sterile soil and liquid medium, both in presence or absence of toluene. We obtained a gapless genome sequence of P. veronii 1YdBTEX2 covering three individual replicons with a total size of 8 Mb, two of which are largely unrelated to current known bacterial replicons. One-hour exposure to toluene, both in soil and liquid, triggered massive transcription (up to 208-fold induction) of multiple gene clusters, such as toluene degradation pathway(s), chemotaxis and toluene efflux pumps. This clearly underlines their key role in the adaptive response to toluene. In comparison to liquid medium, cells in soil drastically changed expression of genes involved in membrane functioning (e.g., lipid composition, lipid metabolism, cell fatty acid synthesis), osmotic stress response (e.g., polyamine or trehalose synthesis, uptake of potassium) and putrescine metabolism, highlighting the immediate response mechanisms of P. veronii 1YdBTEX2 for successful establishment in polluted soil.