CyberKnife Stereotactic Radiosurgery for Uveal Melanoma: First Case Series in Central America.

Introduction: Uveal melanoma is the most common primary intraocular malignancy in adults, affecting primarily the choroid of the eye. Plaque brachytherapy is the most common procedure for the treatment of small choroidal melanoma, especially in posteriorly located tumors. However, modern radiotherapy techniques, such as CyberKnife or Gamma knife stereotactic radiosurgery (SRS) and proton beam radiotherapy, have shown better results in tumor control and eye retention. Recent studies have indicated that SRS is a promising non-invasive, single-session treatment option, with most studies reporting the best outcomes when using ≥21-22 Gy. However, there is no consistent protocol for managing this pathology using CyberKnife, not only in terms of dose but also fractions. Case Presentations: Here, we report the first case series of patients (n = 4, age range 38-64 years, median age 52.5 years) with choroidal UM in Central America who were treated with CyberKnife SRS (22 Gy in one session). During the follow-up (range 25-29 months, median 27.5 months), a 100% control rate with no systemic metastatic disease has been achieved. We found a statistically significant reduction in the largest basal diameter at 24 months for all tumors. However, visual acuity has progressively decreased in most patients. Notably, two of our patients developed radiation maculopathy, and the other two developed radiation retinopathy after SRS. Conclusions: Our findings suggest that future studies should evaluate the use of different prophylactic therapies to prevent the development of side effects. The clinical management of toxicities presented in our report can serve as a reference in the clinical practice of other centers. Our report supports the growing body of evidence showing that CyberKnife radiosurgery is a safe and effective therapeutic option for the treatment of UM.

Pan-cancer analysis of non-oncogene addiction to DNA repair.

Cancer cells usually depend on the aberrant function of one or few driver genes to initiate and promote their malignancy, an attribute known as oncogene addiction. However, cancer cells might become dependent on the normal cellular functions of certain genes that are not oncogenes but ensure cell survival (non-oncogene addiction). The downregulation or silencing of DNA repair genes and the consequent genetic and epigenetic instability is key to promote malignancy, but the activation of the DNA-damage response (DDR) has been shown to become a type of non-oncogene addiction that critically supports tumour survival. In the present study, a systematic evaluation of DNA repair addiction at the pan-cancer level was performed using data derived from The Cancer Dependency Map and The Cancer Genome Atlas (TCGA). From 241 DDR genes, 59 were identified as commonly essential in cancer cell lines. However, large differences were observed in terms of dependency scores in 423 cell lines and transcriptomic alterations across 18 cancer types. Among these 59 commonly essential genes, 14 genes were exclusively associated with better overall patient survival and 19 with worse overall survival. Notably, a specific molecular signature among the latter, characterized by DDR genes like UBE2T, RFC4, POLQ, BRIP1, and H2AFX showing the weakest dependency scores, but significant upregulation was strongly associated with worse survival. The present study supports the existence and importance of non-oncogenic addiction to DNA repair in cancer and may facilitate the identification of prognostic biomarkers and therapeutic opportunities.

The Time for Chronotherapy in Radiation Oncology.

Five decades ago, Franz Halberg conceived the idea of ​​a circadian-based therapy for cancer, given the differential tolerance to treatment derived from the intrinsic host rhythms. Nowadays, different experimental models have demonstrated that both the toxicity and efficacy of several anticancer drugs vary by more than 50% as a function of dosing time. Accordingly, it has been shown that chemotherapeutic regimens optimally timed with the circadian cycle have jointly improved patient outcomes both at the preclinical and clinical levels. Along with chemotherapy, radiation therapy is widely used for cancer treatment, but its effectiveness relies mainly on its ability to damage DNA. Notably, the DNA damage response including DNA repair, DNA damage checkpoints, and apoptosis is gated by the circadian clock. Thus, the therapeutic potential of circadian-based radiotherapy against cancer is mainly dependent upon the control that the molecular clock exerts on DNA repair enzymes across the cell cycle. Unfortunately, the time of treatment administration is not usually considered in clinical practice as it varies along the daytime working hours. Currently, only a few studies have evaluated whether the timing of radiotherapy affects the treatment outcome. Several of these studies show that it is possible to reduce the toxicity of the treatment if it is applied at a specific time range, although with some inconsistencies. In this Perspective, we review the main advances in the field of chronoradiotherapy, the possible causes of the inconsistencies observed in the studies so far and provide some recommendations for future trials.

Functional and Structural Characterization of ClC-1 and Nav1.4 Channels Resulting from CLCN1 and SCN4A Mutations Identified Alone and Coexisting in Myotonic Patients.

Non-dystrophic myotonias have been linked to loss-of-function mutations in the ClC-1 chloride channel or gain-of-function mutations in the Nav1.4 sodium channel. Here, we describe a family with members diagnosed with Thomsen's disease. One novel mutation (p.W322*) in CLCN1 and one undescribed mutation (p.R1463H) in SCN4A are segregating in this family. The CLCN1-p.W322* was also found in an unrelated family, in compound heterozygosity with the known CLCN1-p.G355R mutation. One reported mutation, SCN4A-p.T1313M, was found in a third family. Both CLCN1 mutations exhibited loss-of-function: CLCN1-p.W322* probably leads to a non-viable truncated protein; for CLCN1-p.G355R, we predict structural damage, triggering important steric clashes. The SCN4A-p.R1463H produced a positive shift in the steady-state inactivation increasing window currents and a faster recovery from inactivation. These gain-of-function effects are probably due to a disruption of interaction R1463-D1356, which destabilizes the voltage sensor domain (VSD) IV and increases the flexibility of the S4-S5 linker. Finally, modelling suggested that the p.T1313M induces a strong decrease in protein flexibility on the III-IV linker. This study demonstrates that CLCN1-p.W322* and SCN4A-p.R1463H mutations can act alone or in combination as inducers of myotonia. Their co-segregation highlights the necessity for carrying out deep genetic analysis to provide accurate genetic counseling and management of patients.

Insights into the pathogenicity of missense variants in the forkhead domain of FOX proteins underlying Mendelian disorders.

Forkhead box (FOX) proteins are members of a conserved family of transcription factors. Pathogenic variants in FOX genes have been shown to be responsible for several human genetic diseases. Here, we have studied the molecular and structural features of germline pathogenic variants in seven FOX proteins involved in Mendelian disorders and compared them with those of variants present in the general population (gnomAD). Our study shows that the DNA-binding domain of FOX proteins is particularly sensitive to damaging variation, although some family members show greater mutational tolerance than others. Next, we set to demonstrate that this tolerance depends on the inheritance mode of FOX-linked disorders. Accordingly, genes whose variants underlie recessive conditions are supposed to have a greater tolerance to variation. This is what we found. As expected, variants responsible for disorders with a dominant inheritance pattern show a higher degree of pathogenicity compared to those segregating in the general population. Moreover, we show that pathogenic and likely pathogenic variants tend to affect mutually exclusive sites with respect to those reported in gnomAD. The former also tend to affect sites with lower solvent exposure and a higher degree of conservation. Our results show the value of using publicly available databases and bioinformatics to gain insights into the molecular and structural bases of disease-causing genetic variation.

Mutational survivorship bias: The case of PNKP.

The molecular function of a protein relies on its structure. Understanding how variants alter structure and function in multidomain proteins is key to elucidate the generation of a pathological phenotype. However, one may fall into the logical bias of assessing protein damage only based on the variants that are visible (survivorship bias), which can lead to partial conclusions. This is the case of PNKP, an important nuclear and mitochondrial DNA repair enzyme with both kinase and phosphatase function. Most variants in PNKP are confined to the kinase domain, leading to a pathological spectrum of three apparently distinct clinical entities. Since proteins and domains may have a different tolerability to variation, we evaluated whether variants in PNKP are under survivorship bias. Here, we provide the evidence that supports a higher tolerance in the kinase domain even when all variants reported are deleterious. Instead, the phosphatase domain is less tolerant due to its lower variant rates, a higher degree of sequence conservation, lower dN/dS ratios, and the presence of more disease-propensity hotspots. Together, our results support previous experimental evidence that demonstrated that the phosphatase domain is functionally more necessary and relevant for DNA repair, especially in the context of the development of the central nervous system. Finally, we propose the term "Wald's domain" for future studies analyzing the possible survivorship bias in multidomain proteins.