Most relevant immunohistochemical and clinical data of these series.

<p>NMF:No mutation found; M:Male; F:F; AD:Age at Diagnosis; LDD:Location of the disease at Diagnosis; N:Negative; P:Positive; NV: Not evaluable; HI: High-Intensity; LI:Low-Intensity; L:Low; Mo:Moderate; NP: Not present; NK: Not Known; PI:Perforin Intensity; PG: Perforin Granules; GR: Golgi Region; TC: Troughout the cytoplasm; A: Alive; D:Dead; OS: Overall Survival.</p

An A91V SNV-positive case (A-HE, staining, B-EBER-positive) showing negativity for p53 (C) and low-level of perforin expression (D).

<p>An A91V SNV-negative case (E-HE, staining, F-EBER-positive) showing intense expression of p53 (G) and perforin (H).</p

Mutational landscape of TCL cell lines.

<p>The results of targeted deep sequencing of 16 genes in 20 T-ALL (black), 5 ALCL (dark grey), 3 CTCL (medium grey), 2 NK (light grey), 2 ATLL (diagonal lines) and one T-LGL (dots) cell lines. Mutated genes (rows) are arranged in decreasing order of mutation frequency. Cell lines (columns) are arranged from left to right on the basis of their mutational frequency following gene ranking. HTLV-1-positive cell lines (green) and translocation t(2;5)(p23;q35) (ALK +, dark blue) are showed.</p

Unsupervised hierarchical clustering analysis with 26 immunomarkers.

<p>Each row represents a single cell line; each column represents a single immunomarker. Blue (score 0); white, weak immunostaining (score 1); light red (score 2); red, strong immunoreactivity (score 3); grey, missing data.</p

Mapping of variants in a TCL gene panel.

<p>Schematic of the alterations encoded by SNVs in <i>TP53</i>, <i>NOTCH1</i>, <i>DNMT3A</i>, <i>JAK1</i>, <i>JAK3</i>, <i>STAT3</i> and <i>STAT5B</i>. Type of variation and disease are represented by color and shape, respectively. TAD: transactivation domain; PRD: proline-rich domain; TD: tetramerization domain; C-term: C-terminal domain; HD: heterodimerization domain; TM: transmembrane domain; RAM: Rbp-associated molecule domain; ANK: ankyrin domain; PEST: proline (P), glutamic acid (E), serine (S), threonine (T) degradation domain; ZNF: zinc-finger domain; Mtase: methyltransferase domain.</p

Clinical features, evolution and biological characteristics of the ten CLL patients studied.

<p>Summary of the prognosis factors considered in the clinic and genetic lesions that the ten patients harbor. ND = not done, del = deletion.</p

Mutation validation.

<p>Capillary sequencing chromatograms confirming the seven putative variants. <i>KRAS</i>, <i>SMARCA2</i>, <i>PRKD3</i>, <i>STAT6</i>, <i>LIFR</i> and <i>ILB1</i> showed one point mutation each, depicted as two peaks in the tumoral DNA chromatogram. For <i>NFKBIE</i>, the 4-bp deletion was also confirmed. NT = non-tumoral, TM = tumoral. R = A or G; Y = C or T; S = C or G; K = T or G; /…./ = 4-bp deletion.</p

Validated somatic mutation.

<p>CDS: Coding sequence. NA: not annotated. Chr: chromosome.</p

Schematic representation of the mutated proteins with the primary structural domains highlighted.

<p>Schematic representation of the mutated proteins with the primary structural domains highlighted.</p

PIM kinases as potential therapeutic targets in PTCL.

<p>(A) Gene expression profiling of tumoral samples from 38 human PTCL patients compared with 6 reactive lymph nodes (LN) by microarrays revealed a significantly increased expression of <i>PIM1</i> and <i>PIM2</i> genes (FDR<0.05), but not <i>PIM3</i>. The heatmap is shown in the upper panel, and the relative quantification (Log₂ ratio) comparing PIM expression in PTCL <i>versus</i> LN is shown in the lower panel. (B) GSEA ranked all significantly altered genes between PTCL and LN according to its correlation with <i>PIM1</i> or <i>PIM2</i> expression and displayed them in the red-to-blue bar. Each gene belonging to every pathway was interrogated whether it appeared positively (in the red region of the bar) or negatively (in the blue side) correlated. Using this approach GSEA identified a positive and significant correlation between <i>PIM1</i> and <i>PIM2</i> expression and Jak/STAT, NF-κB and IL-2 signaling pathways in the PTCL molecular signature (FDR<0.25). (C) PIM family genes mRNA level was measured by RT-qPCR in eight PTCL cell lines and (D) primary tumoral T cells from 5 Sézary Syndrome patients (SS #1–5), and compared with normal T cells isolated from 3 healthy donors (Control #1–3). The relative RNA amount of PIM has been calculated as a relative quantification, as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0112148#s2" target="_blank">Methods</a> section (RQ = 2^−ΔCt), normalized with non-tumoral cells: RQ in PTCL/RQ in healthy #3. In both settings, <i>PIM1</i>, and especially <i>PIM2</i>, but not <i>PIM3</i> expression was found to be increased in PTCL. (E) PIM kinase protein basal levels in PTCL cell lines measured by Western blot. PIM1 and PIM2 isoforms are also shown. (F) Distribution of PIM2 protein in a series of tumoral samples from 136 PTCL patients measured by immunohistochemistry. Negative, weakly positive and strongly positive samples were defined by the presence of <5%, 5–20% and >20% positive cells. (G) Distribution of PIM2 protein in the most common PTCL subtypes measured by immunohistochemistry.</p