Multiple mutations at exon 2 of RHOA detected in plasma from patients with peripheral T-cell lymphoma.

The mutational landscape of peripheral T-cell lymphoma (PTCL) is being revealed through sequencing of lymph node samples, but there has been little work on the mutational load that is present in cell-free DNA (cfDNA) from plasma. We report targeted sequencing of cfDNA from PTCL patients to demonstrate c.50G>T (p.Gly17Val) in RHOA as previously described in angioimmunoblastic T-cell lymphoma (AITL) and a group of PTCL not otherwise specified (NOS) but also detect novel mutations at c.73A>G (p.Phe25Leu) and c.48A>T (p.Cys16*) of exon 2, which were confirmed by Sanger sequencing. In a group of AITL and PTCL-NOS analyzed by droplet digital polymerase chain reaction, 63% (12/19) showed c.50G>T (p.Gly17Val), 53% (10/19) c.73A>G (p.Phe25Leu), and 37% (7/19) c.48A>T (pCys16*). Sequencing of lymph node tissue in 3 out of 9 cases confirmed the presence of c.73A>G (p.Phe25Leu). Inspection of individual sequencing reads from individual patients showed that a single RHOA allele could contain >1 mutation, suggesting haplotypes of mutations at RHOA. Serial sampling showed changes to RHOA mutational frequency with treatment and the apparent occurrence of clones bearing specific haplotypes associated with relapse. Therefore, sequencing of RHOA from cfDNA has revealed new mutations and haplotypes. The clinical significance of these findings will need to be explored in clinical trials, but liquid biopsy might have potential for guiding treatment decisions in PTCL.

Shprintzen-Goldberg Syndrome: A Rare Disorder.

The Shprintzen-Goldberg syndrome (SGS) or velo-cardio-facial syndrome (VCFS) is an extremely rare disorder of connective tissue with a characteristic facial dysmorphism, marfanoid features, craniosynostosis, dolichocephaly, cardiovascular anomalies and mild to moderate mental retardation. It may be a de novo gene mutation or inherited as an autosomal dominant disorder having SKI gene and Fibrillin-1 gene (FBN1) mutations, located on chromosome 15q21.1. We report a case of a 3-month, developmentally delayed male infant admitted to the hospital with syndromic facies, craniosynostosis, joint laxity and on echocardiography, aortic root dilatation. A probable diagnosis of SGS was made on the clinical grounds. We did not have the facility for genetic chromosomal analysis. He was discharged with family counselling and follow-up for future developmental rehabilitation.

Amniotic Fluid Embolism.

Amniotic fluid embolism is a rare and severe problem in obstetric patients. We experienced a 21-year primigravida who underwent emergency cesarean section due to sudden collapse and fetal distress after rupture of membranes in labour. Subsequently, she developed intraoperative coagulopathy, hemorrhage, hypotension, and respiratory collapse requiring ventilation. Both maternal and neonatal lives were saved with full recovery and discharged in stable condition. The clinical diagnosis of amniotic fluid embolism (AFE) was made, which is a very rare complication of pregnancy/puerperium with varying presentation, ranging from cardiac arrest, circulatory collapse and death through mild degrees of organ system dysfunction with or without coagulopathy. AFE has no definitive test for its diagnosis; the detection of fetal elements in the maternal vasculature is non-specific. The treatment is largely supportive and there is no specific therapy. However, mortality of this condition remains high, if not timely intervened.

PD1hi cells associate with clusters of proliferating B-cells in marginal zone lymphoma.

BACKGROUND: Abnormally sustained immune reactions drive B-cell proliferation in some cases of marginal zone lymphoma but the CD4+ T-cell subsets, which are likely to contribute to the B-cell responses in the tumour microenvironment, are not well characterised and neither has the spatial distribution of the different subsets in involved lymph nodes been investigated. METHODS: Employing a workflow of multiplex semi-automated immunohistochemistry combined with image processing we investigated association between infiltrating T-cells and proliferating lymphoma B-cells. RESULTS: Both total numbers of activating follicular helper (Tfh) cells (defined by high expression of PD1) and suppressive regulatory (Treg) T-cells (defined by FOXP3+ expression) and the Tfh:Treg ratio, assessed over relatively large areas of tissue, varied among cases of marginal zone lymphoma. We determined spatial distribution and demonstrated that PD1hi cells showed significantly more clustering than did FOXP3+. To investigate the association of infiltrating T-cells with lymphoma B-cells we employed Pearson correlation and Morisita-Horn index, statistical measures of interaction. We demonstrated that PD1hi cells were associated with proliferating B-cells and confirmed this by nearest neighbour analysis. CONCLUSIONS: The unexpected architectural complexity of T-cell infiltration in marginal zone lymphoma, revealed in this study, further supports a key role for Tfh cells in driving proliferation of lymphoma B-cells. We demonstrate the feasibility of digital analysis of spatial architecture of T-cells within marginal zone lymphoma and future studies will be needed to determine the clinical importance of these observations.