BRAFV600E mutation positive metastatic melanoma in a young woman treated with anti-BRAF/anti MEK combination: a case report

The recent Combi-v [1] and Combi-d [2,3], phase III randomized trials, showed, respectively, an OS benefit with Dabrafenib/Trametinib combination versus Vemurafenib and an improvement in PFS and ORR with the same combination versus Dabrafenib alone, in BRAF V600E/K mutation positive metastatic or unresectable cutaneous melanoma. We report the case of a young patient with metastatic melanoma treated with anti- BRAF/antiMEK combination

T-cell specific deregulation of Notch3 receptor induces alterations in the development of myeloid compartment unveiled by the deletion of NF-kappaB/p50 expression

Purpose/Objective: T-cell specific deregulation of Notch3 in transgenicmice (N3-tg), induces the development of a T-cell acute lymphoblastic leukemia (T-ALL), sustained by the constitutive activation of NF-kB canonical pathway. Besides, Notch signalling modulation in bone-marrow stromal cells or in hematopoietic stem cells, has been related to alterations in differentiation/proliferation processes of myeloid cells. To clarify the Notch/NF-kB relationships in the progression of T-ALL and the effects of a T-cell specific deregulation of Notch on myeloid compartment, we decided to delete NF-kB canonical pathway in N3-tg mice. Materials and methods: We generated N3-tg/p50-/- mice, deleted of the NF-kB/p50 subunit in a Notch3 transgenic background. The follow-up of double mutant versus N3-tg mice versus relative controls was conducted and immunophenotyping of hematopoietic cell subsets was performed at different age and in multiple tissues from the indicated animals by flow-cytometry tecniques. Total RNA and protein extract samples, derived from sorted T- or myeloid-cells of our mice models, were processed for RT-qPCR and Western blotting analysis, respectively, to test the expression of Notch-related molecules. Results: The progression of T-ALL, as defined by the peripheral expansion of immature CD4+ CD8+ T cells, was strongly inhibited in N3-tg/p50-/- versus N3-tg mice. However, the double mutant mice succumb earlier than N3-tg counterparts displaying a dramatic increase of Mac1+ Gr1+ myeloid cells in both spleen and blood, as well as of granulocyte/monocyte progenitors in the bone marrow. The expansion of myeloid subsets was detectable at a lower extent also in N3-tg versus wild-type mice. Preliminary data indicate that Mac1+ Gr1+ cells do not express Notch3, suggesting that this receptor may influence the equilibrium of the myeloid compartment mainly in trans, possibly through its interaction with the Jagged-1 ligand. Conclusions: Our results suggest that the NF-kB canonical pathway deletion inhibits the T-ALL progression, thus unveiling the influence of Notch signalling modulation on the behaviour of myeloid cells. We provide a useful model to extend our understanding of Notch/NF-kB interplay in driving the relationships between lymphoid and myeloid compartments in the context of hematological malignancy

The Outcome of Notch3-dependent T Cell Leukemia Is Modified By NF-kappaB Deletion

Background: The Notch3 deregulation inside T-cell compartment of transgenic (N3-tg) mice, induces an aggressive form of T-cell acute lymphoblastic leukemia (T-ALL), strongly sustained by an NF-κB constitutive activation, mainly represented by the p50/p65-dependent canonical pathway. To clarify the Notch/NF-κB relationships in the onset/progression of T-ALL, we decided to inhibit NF-κB canonical pathway in N3-tg mice. Methods: We generated N3-tg/p50-/- mice, deleted of the NF-κB/p50 subunit in a Notch3 transgenic background. The follow-up of N3-tg/p50-/- versus N3- tg mice was conducted and hematopoietic cell analysis was performed at different ages and in multiple tissues from the indicated animals by flow-cytometry techniques. Results: The p50 deletion inhibited the progression of T-ALL in N3-tg/p50-/- mice, as defined primarily by the peripheral expansion of immature CD4+CD8+ T cells. Surprisingly, the double mutant mice succumb earlier than N3-tg counterparts. Moribund N3-tg/p50-/- mice display the trait of a myeloproliferative disease, with the dramatic expansion of Mac1+Gr1+ myeloid cells in both spleen and blood, as well as of granulocyte/monocyte progenitors in the bone marrow. Preliminary data indicate that these cells do not express Notch3, suggesting that in the absence of p50 expression, Notch3 is able to mainly influence the equilibrium of the myeloid compartment in trans. Conclusions: The results presented suggest that the ablation of NF-κB canonical pathway may strongly impact on the outcomes of a T cell specific deregulation of Notch signaling. Thus, providing a useful experimental model to extend our understanding of Notch/NF-κB interplay and to unravel novel strategies for the therapy of different hematological malignancies

Deletion of NF-kappa/p50 influences the outcome of Notch3-dependent T cell leukemia

The deregulation of Notch3 signaling inside T-cell compartment of transgenic (N3-tg) mice, induces an aggressive form of T-cell acute lymphoblastic lymphoma (T-ALL), sustained by a constitutive activation of the NF-B canonical pathway, mainly represented by the p50/p65 heterodimer. To clarify the Notch/NF-B relationships in the development of T-ALL, we generated double mutant mice, deleted of the NF-B/p50 subunit in a Notch3 transgenic background (N3-tg/p50-/-). The follow-up of N3-tg/p50-/- versus N3-tg mice revealed that p50 deletion strongly inhibits the development of Notch3-dependent T-ALL. Surprisingly, double mutant succumb earlier than N3-tg mice, displaying the trait of a myeloproliferative disease, with an aberrant accumulation of Mac1+Gr1+ myeloid cells in both spleen and peripheral blood, as well as of granulocyte/monocyte progenitors in the bone marrow. Our preliminary results suggest that Notch3 overexpression in T-cell compartment is able to influence, possibly in trans, the equilibrium of the myeloid compartment and that the ablation of NF-B canonical pathway may impact on the outcomes of a T-cell specific deregulation of Notch signaling. We provide a useful experimental model to extend our understanding of Notch/NF-B interplay in hemopoietic system and to unravel novel relationships between lymphoid and myeloid differentiation

Cutaneous squamous cell carcinoma: from pathophysiology to novel therapeutic approaches

: Cutaneous squamous cell carcinoma (cSCC), a non-melanoma skin cancer, is a keratinocyte carcinoma representing one of the most common cancers with an increasing incidence. cSCC could be in situ (e.g., Bowen's disease) or an invasive form. A significant cSCC risk factor is advanced age, together with cumulative sun exposure, fair skin, prolonged immunosuppression, and previous skin cancer diagnoses. Although most cSCCs can be treated by surgery, a fraction of them recur and metastasize, leading to death. cSCC could arise de novo or be the result of a progression of the actinic keratosis, an in situ carcinoma. The multistage process of cSCC development and progression is characterized by mutations in the genes involved in epidermal homeostasis and by several alterations, such as epigenetic modifications, viral infections, or microenvironmental changes. Thus, cSCC development is a gradual process with several histological- and pathological-defined stages. Dermoscopy and reflectance confocal microscopy enhanced the diagnostic accuracy of cSCC. Surgical excision is the first-line treatment for invasive cSCC. Moreover, radiotherapy may be considered as a primary treatment in patients not candidates for surgery. Extensive studies of cSCC pathogenic mechanisms identified several pharmaceutical targets and allowed the development of new systemic therapies, including immunotherapy with immune checkpoint inhibitors, such as Cemiplimab, and epidermal growth factor receptor inhibitors for metastatic and locally advanced cSCC. Furthermore, the implementation of prevention measures has been useful in patient management

Mouse Sertoli Cells Sustain De Novo Generation of Regulatory T Cells by Triggering the Notch Pathway Through Soluble JAGGED1

FOXP3(+) regulatory T cells (Tregs) are central to the maintenance of immunological homeostasis and tolerance. It has long been known that Sertoli cells are endowed with immune suppressive properties; however, the underlying mechanisms as well as the effective nature and role of soluble factors secreted by Sertoli cells have not been fully elucidated as yet. We hypothesized that conditioned medium from primary mouse Sertoli cells (SCCM) may be able and sufficient to induce Tregs. By culturing CD4(+)CD25(-)EGFP(-) T splenocytes purified from FOXP3-EGFP knock-in mice in SCCM, here we show, by flow cytometry and suppression assay, the conversion of peripheral CD4(+)FOXP3(-) T cells into functional CD4(+)FOXP3(+) Tregs. We also demonstrate that the Notch/Jagged1 axis is involved in regulating the de novo generation of Tregs although this process is transforming growth factor-beta1 (TGF-B) dependent. In particular, we identified by Western blot analysis a soluble form of JAGGED1 (JAG1) in SCCM that significantly influences the induction of Tregs, as demonstrated by performing the conversion assay in presence of a JAG1-specific neutralizing antibody. In addition, we show that SCCM modulates the Notch pathway in converted Tregs by triggering the recruitment of the Notch-specific transcription factor CSL/RBP-Jk to the Foxp3 promoter and by inducing the Notch target gene Hey1, as shown by chromatin immunoprecipitation assay and by real time-RT-PCR experiments, respectively. Overall, these results contribute to a better understanding of the molecular mechanisms involved in Sertoli cell-mediated immune tolerance and provide a novel approach to generate ex vivo functional Tregs for therapeutic purpose

Cutaneous Squamous Cell Carcinoma: From Pathophysiology to Novel Therapeutic Approaches

Cutaneous squamous cell carcinoma (cSCC), a non-melanoma skin cancer, is a keratinocyte carcinoma representing one of the most common cancers with an increasing incidence. cSCC could be in situ (e.g., Bowen’s disease) or an invasive form. A significant cSCC risk factor is advanced age, together with cumulative sun exposure, fair skin, prolonged immunosuppression, and previous skin cancer diagnoses. Although most cSCCs can be treated by surgery, a fraction of them recur and metastasize, leading to death. cSCC could arise de novo or be the result of a progression of the actinic keratosis, an in situ carcinoma. The multistage process of cSCC development and progression is characterized by mutations in the genes involved in epidermal homeostasis and by several alterations, such as epigenetic modifications, viral infections, or microenvironmental changes. Thus, cSCC development is a gradual process with several histological- and pathological-defined stages. Dermoscopy and reflectance confocal microscopy enhanced the diagnostic accuracy of cSCC. Surgical excision is the first-line treatment for invasive cSCC. Moreover, radiotherapy may be considered as a primary treatment in patients not candidates for surgery. Extensive studies of cSCC pathogenic mechanisms identified several pharmaceutical targets and allowed the development of new systemic therapies, including immunotherapy with immune checkpoint inhibitors, such as Cemiplimab, and epidermal growth factor receptor inhibitors for metastatic and locally advanced cSCC. Furthermore, the implementation of prevention measures has been useful in patient management

The Impact of Drug–Drug Interactions on the Toxicity Profile of Combined Treatment with BRAF and MEK Inhibitors in Patients with BRAF-Mutated Metastatic Melanoma

Background: BRAF and MEK inhibition is a successful strategy in managing BRAF-mutant melanoma, even if the treatment-related toxicity is substantial. We analyzed the role of drug–drug interactions (DDI) on the toxicity profile of anti-BRAF/anti-MEK therapy. Methods: In this multicenter, observational, and retrospective study, DDIs were assessed using Drug-PIN software (V 2/23). The association between the Drug-PIN continuous score or the Drug-PIN traffic light and the occurrence of treatment-related toxicities and oncological outcomes was evaluated. Results: In total, 177 patients with advanced BRAF-mutated melanoma undergoing BRAF/MEK targeted therapy were included. All grade toxicity was registered in 79% of patients. Cardiovascular toxicities occurred in 31 patients (17.5%). Further, 94 (55.9%) patients had comorbidities requiring specific pharmacological treatments. The median Drug-PIN score significantly increased when the target combination was added to the patient’s home therapy (p-value p-value = 0.048). The Drug-PIN traffic light (p = 0.00821) and the Drug-PIN score (p = 0.0291) were seen to be significant predictors of cardiotoxicity. Patients with low-grade vs. high-grade interactions showed a better prognosis regarding overall survival (OS) (p = 0.0045) and progression-free survival (PFS) (p = 0.012). The survival analysis of the subgroup of patients with cardiological toxicity demonstrated that patients with low-grade vs. high-grade DDIs had better outcomes in terms of OS (p = 0.0012) and a trend toward significance in PFS (p = 0.068). Conclusions: DDIs emerged as a critical issue for the risk of treatment-related cardiovascular toxicity. Our findings support the utility of DDI assessment in melanoma patients treated with BRAF/MEK inhibitors