Surgery Alone (Without Adjuvant Radiation) Adequately Treats Histologic Perineural Basal Cell Carcinomas: A Systematic Review With Meta-Analysis.

BACKGROUND: Histologic perineural invasion (PNI) in basal cell carcinomas (BCC) lacks evidence-based treatment guidelines. OBJECTIVE: Systematically review and analyze treatment outcomes of BCC with histologic PNI (PNBCC). MATERIALS AND METHODS: PubMed, Embase, and Cochrane Reviews were searched through June 25, 2021. Thirteen eligible cohort studies were meta-analyzed. RESULTS: 502 of 713 PNBCC were treated with Mohs Surgery (MMS), wide local excision (WLE), or surgery (MMS or WLE) with adjuvant radiation (Surg + RT). Overall 5-year local control (LC) was 97.2% and cancer-specific survival (CSS) was 99.6%. Surg and Surg + RT did not differ in recurrence (2.1% vs 4.7%; p-value 0.56; RR 1.51 [0.37, 6.20]), LC (97.9% vs 96.2%; p-value 0.19; RR 0.98 [0.96, 1.01]) or CSS (100% vs 99.1%; p-value 0.40; RR 0.99 [0.95, 1.02]). LIMITATIONS: No randomized controlled trials were found. Outcome data were often lacking. CONCLUSION: Overall LC and CSS were high at median 5-year follow-up for surgery alone and Surg + RT. Surgery alone and Surg + RT demonstrated statistically equivalent outcomes. We do not recommend adjuvant radiation therapy for solely histologic PNBCC if clear margins are achieved.

Cutting Edge: Quantitative Determination of CD40L Threshold for IL-12 and IL-23 Production from Dendritic Cells.

Early secretion of IL-12 by mouse dendritic cells (DCs) instructs T cells to make IFN-γ. However, only activated, but not naive T cells are able to license DCs for IL-12 production. We hypothesized that it might be due to different levels of CD40L expression on the surface of these cells, as CD40 signals are required for IL-12 production. Using quantitative cell-free systems incorporating CD40L in lipid bilayers combined with total internal reflection fluorescence microscopy and flow cytometry, we show that as low as ∼200 CD40L molecules/µm2 in combination with IL-4 is sufficient to induce IL-12 production by DCs. Remarkably, CD40L alone is adequate to induce IL-23 secretion by DCs. Thus, although activated T cells have somewhat higher levels of CD40L, it is the combination of CD40L and the cytokines they secrete that licenses DCs and influences the effector class of the immune response.

Essential role of ubiquitin and TSG101 protein in formation and function of the central supramolecular activation cluster.

Agonist MHC-peptide complexes in the immunological synapse (IS) signal through T cell receptor (TCR) microclusters (MCs) that converge into a central supramolecular activation cluster (cSMAC). The determinants and function of the cSMAC remain unknown. We demonstrate an essential role for ubiquitin (Ub) and TSG101, but less so for HRS, in signal processing events at the cSMAC. Using siRNA in primary T cells, we show that Ub recognition by TSG101 is required for cSMAC formation, TCR MC signal termination, TCR downregulation, and segregation of TCR-MHC-peptide from PKC-theta-enriched signaling complexes. Weak agonist MHC-peptide induced CD80-dependent TCR MCs that dissociated in the center of the IS without recruiting TSG101. These results support TSG101-dependent recognition of CD80-independent TCR MCs as a molecular checkpoint for TCR downregulation.

Diffusion and signaling revisited.

In this issue of Immunity, Andrews et al. (2009) used single-molecule fluorescence microscopy to demonstrate that the IgE receptor FcepsilonRI in the plasma membrane can signal in a mobile state.

Kinetics of early T cell receptor signaling regulate the pathway of lytic granule delivery to the secretory domain.

Cytolytic granules mediate killing of virus-infected cells by cytotoxic T lymphocytes. We show here that the granules can take long or short paths to the secretory domain. Both paths utilized the same intracellular molecular events, which have different spatial and temporal arrangements and are regulated by the kinetics of Ca(2+)-mediated signaling. Rapid signaling caused swift granule concentration near the microtubule-organizing center (MTOC) and subsequent delivery by the polarized MTOC directly to the secretory domain-the shortest path. Indolent signaling led to late recruitment of granules that moved along microtubules to the periphery of the synapse and then moved tangentially to fuse at the outer edge of the secretory domain-a longer path. The short pathway is associated with faster granule release and more efficient killing than the long pathway. Thus, the kinetics of early signaling regulates the quality of the T cell cytolytic response.

Observation of Dog-Ear Regression by Anatomical Location.

BACKGROUND: When an excision is performed by a method other than elliptical excision, direct primary wound closure can result in standing cones or "dog-ears." In 2008, Lee and colleagues noted that dog-ears of <8 mm in height have a statistically greater tendency to resolve without further surgical correction than larger dog-ears. OBJECTIVE: To stratify dog-ears by anatomic location and inform on the need for correction at the time of surgery. MATERIALS AND METHODS: After tumor extirpation, patients were counseled that primary closure of the surgical wound would result in dog-ears at the wound apices. Dog-ears were left uncorrected in participating patients. At 6 months, patients were assessed for resolution of the dog-ears and asked to rate the appearance of the scar. RESULTS: A total of 140 dog-ears were observed in the study period. Anatomical locations included the hand/foot, trunk, limb, and head/neck. Among these dog-ears, 114/140 (81%) showed complete resolution. Patient satisfaction with the scar appearance correlated well with the dog-ear resolution, with most patients rating the appearance of the scar as good to excellent. CONCLUSION: This study suggests that dog-ears on the hand and dog-ears ≤4 mm on the trunk may be observed without any final cosmetic penalty.

TCR Microclusters pre-exist and contain molecules necessary for TCR signal transduction.

TCR-dependent signaling events have been observed to occur in TCR microclusters. We found that some TCR microclusters are present in unstimulated murine T cells, indicating that the mechanisms leading to microcluster formation do not require ligand binding. These pre-existing microclusters increase in absolute number following engagement by low-potency ligands. This increase is accompanied by an increase in cell spreading, with the result that the density of TCR microclusters on the surface of the T cell is not a strong function of ligand potency. In characterizing their composition, we observed a constant number of TCRs in a microcluster, constitutive exclusion of the phosphatase CD45, and preassociation with the signaling adapters linker for activation of T cells and growth factor receptor-bound protein 2. The existence of TCR microclusters prior to ligand binding in a state that is conducive for the initiation of downstream signaling could explain, in part, the rapid kinetics with which TCR signal transduction occurs.

Utility of Wood's light in margin determination of melanoma in situ after excisional biopsy.

BACKGROUND: Margin evaluation of melanoma in situ (MIS) is difficult because of its ill-defined clinical borders. Wood's light examination is commonly used to help delineate MIS margin before excision. OBJECTIVE: To prospectively study the accuracy of preoperative Wood's light examination for margin assessment of MIS. MATERIALS AND METHODS: The authors evaluated 60 patients before excision of MIS under white light and Wood's light. Staged excision was performed using the square procedure technique. After achieving clear margins, they compared final wound size with expected wound size if surgical margins had been based on Wood's light examination. RESULTS: Seven patients (11.7%) had Wood's light enhancement beyond the visible margin of the biopsy site. In all 7, increased wounding would have occurred if the surgical margins had been based on Wood's light examination. In 1 of the 7, use of the Wood's light examination would have reduced the surgical stages needed by 1 stage but would have increased the wound size by 83.3%. CONCLUSION: Wood's light examination has limited utility if complete excisional biopsy of MIS is performed before treatment. In this study, surgical margin based on the Wood's light examination would have resulted in an increased average wound size and would not have reduced the number of stages needed when performing the square procedure.

Endogenous viral antigen processing generates peptide-specific MHC class I cell-surface clusters.

Sensitivity is essential in CD8+ T-cell killing of virus-infected cells and tumor cells. Although the affinity of the T-cell receptor (TCR) for antigen is relatively low, the avidity of T cell-antigen-presenting cell interactions is greatly enhanced by increasing the valence of the interaction. It is known that TCRs cluster into protein islands after engaging their cognate antigen (peptides bound to MHC molecules). Here, we show that mouse K(b) class I molecules segregate into preformed, long-lasting (hours) clusters on the antigen-presenting cell surface based on their bound viral peptide. Peptide-specific K(b) clustering occurs when source antigens are expressed by vaccinia or vesicular stomatitis virus, either as proteasome-liberated precursors or free intracellular peptides. By contrast, K(b)-peptide complexes generated by incubating cells with synthetic peptides are extensively intermingled on the cell surface. Peptide-specific complex sorting is first detected in the Golgi complex, and compromised by removing the K(b) cytoplasmic tail. Peptide-specific clustering is associated with increased T-cell sensitivity: on a per-complex basis, endogenous SIINFEKL activates T cells more efficiently than synthetic SIINFEKL, and wild-type K(b) presents endogenous SIINFEKL more efficiently than tailless K(b). We propose that endogenous processing generates peptide-specific clusters of class I molecules to maximize the sensitivity and speed of T-cell immunosurveillance.

Ligand-mediated friction determines morphodynamics of spreading T cells.

Spreading of T cells on antigen presenting cells is a crucial initial step in immune response. Spreading occurs through rapid morphological changes concomitant with the reorganization of surface receptors and of the cytoskeleton. Ligand mobility and frictional coupling of receptors to the cytoskeleton were separately recognized as important factors but a systematic study to explore their biophysical role in spreading was hitherto missing. To explore the impact of ligand mobility, we prepared chemically identical substrates on which molecules of anti-CD3 (capable of binding and activating the T cell receptor complex), were either immobilized or able to diffuse. We quantified the T cell spreading area and cell edge dynamics using quantitative reflection interference contrast microscopy, and imaged the actin distribution. On mobile ligands, as compared to fixed ligands, the cells spread much less, the actin is centrally, rather than peripherally distributed and the edge dynamics is largely altered. Blocking myosin-II or adding molecules of ICAM1 on the substrate largely abrogates these differences. We explain these observations by building a model based on the balance of forces between activation-dependent actin polymerization and actomyosin-generated tension on one hand, and on the frictional coupling of the ligand-receptor complexes with the actin cytoskeleton, the membrane and the substrate, on the other hand. Introducing the measured edge velocities in the model, we estimate the coefficient of frictional coupling between T Cell receptors or LFA-1 and the actin cytoskeleton. Our results provide for the first time, to our knowledge, a quantitative framework bridging T cell-specific biology with concepts developed for integrin-based mechanisms of spreading.

IL-15/IL-15Rα-Fc fusion protein XmAb24306 potentiates activity of CD3 bispecific antibodies through enhancing T cell expansion.

Insufficient quantity of functional T cells is a likely factor limiting clinical activity of T cell bispecific antibodies, especially in solid tumor indications. We hypothesized that XmAb24306 (efbalropendekin alfa), a lymphoproliferative interleukin (IL)-15/IL-15 receptor α (IL-15Rα) Fc-fusion protein, may potentiate the activity of T cell dependent (TDB) antibodies. Activation of human peripheral T cells by cevostamab, an anti-FcRH5/CD3 TDB, or anti-HER2/CD3 TDB resulted in upregulation of IL-2/15Rß (CD122) receptor subunit in nearly all CD8+ and majority of CD4+ T cells, suggesting that TDB treatment may sensitize T cells to the IL-15. XmAb24306 enhanced T cell bispecific antibody induced CD8+ and CD4+ T cell proliferation and expansion. In vitro combination of XmAb24306 with cevostamab or anti-HER2/CD3 TDB resulted in significant enhancement of tumor cell killing, which was reversed when T cell numbers were normalized, suggesting that T cell expansion is the main mechanism for the observed benefit. Pre-treatment of immune competent mice with a mouse-reactive surrogate of XmAb24306 (mIL-15-Fc) resulted in significant increase of T cells in blood, spleen and in tumors and converted transient anti-HER2/CD3 TDB responses to complete durable responses. In summary, our results support the hypothesis where the number of tumor infiltrating T cells is rate limiting for the activity of solid tumor targeting TDBs. Upregulation of CD122 by TDB treatment and the observed synergy with XmAb24306 and T cell bispecific antibodies supports clinical evaluation of this novel immunotherapy combination.

Complex PK-PD of an engineered IL-15/IL-15Rα-Fc fusion protein in cynomolgus monkeys: QSP modeling of lymphocyte dynamics.

XmAb24306 is a lymphoproliferative interleukin (IL)-15/IL-15 receptor α (IL-15Rα) Fc-fusion protein currently under clinical investigation as an immunotherapeutic agent for cancer treatment. XmAb24306 contains mutations in IL-15 that attenuate its affinity to the heterodimeric IL-15 receptor ßγ (IL-15R). We observe substantially prolonged pharmacokinetics (PK) (half-life ∼ 2.5 to 4.5 days) in single- and repeat-dose cynomolgus monkey (cyno) studies compared to wild-type IL-15 (half-life ∼ 1 hour), leading to increased exposure and enhanced and durable expansion of NK cells, CD8+ T cells and CD4-CD8- (double negative [DN]) T cells. Drug clearance varied with dose level and time post-dose, and PK exposure decreased upon repeated dosing, which we attribute to increased target-mediated drug disposition (TMDD) resulting from drug-induced lymphocyte expansion (i.e., pharmacodynamic (PD)-enhanced TMDD). We developed a quantitative systems pharmacology (QSP) model to quantify the complex PKPD behaviors due to the interactions of XmAb24306 with multiple cell types (CD8+, CD4+, DN T cells, and NK cells) in the peripheral blood (PB) and lymphoid tissues. The model, which includes nonspecific drug clearance, binding to and TMDD by IL15R differentially expressed on lymphocyte subsets, and resultant lymphocyte margination/migration out of PB, expansion in lymphoid tissues, and redistribution to the blood, successfully describes the systemic PK and lymphocyte kinetics observed in the cyno studies. Results suggest that after 3 doses of every-two-week (Q2W) doses up to 70 days, the relative contributions of each elimination pathway to XmAb24306 clearance are: DN T cells > NK cells > CD8+ T cells > nonspecific clearance > CD4+ T cells. Modeling suggests that observed cellular expansion in blood results from the influx of cells expanded by the drug in lymphoid tissues. The model is used to predict lymphoid tissue expansion and to simulate PK-PD for different dose regimens. Thus, the model provides insight into the mechanisms underlying the observed PK-PD behavior of an engineered cytokine and can serve as a framework for the rapid integration and analysis of data that emerges from ongoing clinical studies in cancer patients as single-agent or given in combination.

A new fractionation assay, based on the size of formaldehyde-crosslinked, mildly sheared chromatin, delineates the chromatin structure at promoter regions.

To explore the higher order structure of transcribable chromatin in vivo, its local configuration was assessed through the accessibility of the chromatin to crosslinking with formaldehyde. The application of crosslinked and mildly sheared chromatin to sedimentation velocity centrifugation followed by size-fractionation of the DNA enabled us to biochemically distinguish between chromatin with heavily versus sparsely crosslinkable structures. The separated fractions showed a good correlation with gene expression profiles. Genes with poor crosslinking around the promoter region were actively transcribed, while transcripts were hardly detected from genes with extensive crosslinking in their promoter regions. For the inducible gene, Il2, the distribution of the promoter shifted in the gradient following T-cell receptor stimulation, consistent with a change in structure at this locus during activation. The kinetics of this switch preceded the chromatin change observed in a DNase I accessibility assay. Thus, this new chromatin fractionation technique has revealed a change in chromatin structure that has not been previously characterized.

Extramedullary Tibial Guide Alignment Is Not Affected by Excess Lower Limb Fat Distribution in Total Knee Arthroplasty.

Background The goal of this study is to investigate whether excess lower limb fat distribution affects tibial guide alignment in conventional total knee arthroplasty (TKA) with extramedullary guides. A thicker soft tissue envelope may affect the accuracy of extramedullary cutting guide placement and subsequent instrumentation. Previous studies have used body mass index (BMI) to stratify patients, a poor proxy of lower limb fat distribution, which may explain conflicting results reported on this topic to date. This study overcomes this issue by using a novel, radiographic anthropometric index to assess lower limb fat distribution. Methodology This is a single-surgeon, single-implant, single-centre retrospective series of 102 consecutive primary TKAs. The suprapatellar fat index (SPFI) and BMI were recorded for all patients, and postoperative tibial component alignment measurements were calculated. Secondary outcome measures included femoral component alignment, femorotibial alignment, length of hospital stay, tourniquet time, blood loss, and complications/reoperations. Results In this study, 102 patients (average age of 69) had an average BMI of 30.8 kg/m2 (19.2-45.5 kg/m2) and an average SPFI of 0.26 (0.09-0.57). Multiple regression analysis demonstrated that increasing leg fat distribution did not affect tibial component alignment in the coronal or sagittal plane. Conclusions Excess lower limb fat distribution, simply measured using the SPFI, does nothave a significant effect on tibial component positioning when extramedullary guides are used in conventional TKA.

The management of combined ACL and MCL injuries: A systematic review.

Background: The management of combined anterior cruciate ligament (ACL) and medial collateral ligament (MCL) injuries remains contentious. Clinical outcomes of surgical, conservative, and combined approaches have been described in a range of prospective and retrospective studies. The aim of the current systematic review was to evaluate these outcomes and assess the study methodologies. Methods: A comprehensive literature search of the following databases was performed: PubMed, OVID, Cochrane Database of Systematic Reviews and Google Scholar. Studies were assessed using the Coleman Methodology Score. Results: 52 articles were included (3 randomised controlled trials, 8 prospective comparative studies, 17 retrospective comparative studies and 24 case series). Outcome measures were heterogeneous amongst articles. The most common outcomes assessed were AP laxity, Lysholm score and medial/valgus laxity. Complications at varying follow-up times with differing grades of MCL injury were reported in 25 (48%) studies. Evidence was conflicting, with no consensus from the available published literature regarding the best method of treatment for a combined ACL and MCL injury. Conclusions: Heterogeneous outcome measures and limited randomised controlled trials prevent advocacy of a single treatment option. Good outcomes have been reported from repair, reconstruction and conservative management of the MCL together with ACL reconstruction. Further prospective comparative data is required to evaluate MCL management choice and prognostic signs for successful nonsurgical MCL treatment.

Actin and agonist MHC-peptide complex-dependent T cell receptor microclusters as scaffolds for signaling.

T cell receptor (TCR) microclusters form within seconds of T cell contact with supported planar bilayers containing intercellular adhesion molecule-1 and agonist major histocompatibility complex (MHC)-peptide complexes, and elevation of cytoplasmic Ca2+ is observed within seconds of the first detectable microclusters. At 0-30 s after contact, TCR microclusters are colocalized with activated forms of Lck, ZAP-70, and the linker for activation of T cells. By 2 min, activated kinases are reduced in the older central microclusters, but are abundant in younger peripheral microclusters. By 5 min, TCR in the central supramolecular activation cluster have reduced activated kinases, whereas faint peripheral TCR microclusters efficiently generated activated Lck and ZAP-70. TCR microcluster formation is resistant to inhibition by Src family kinase inhibitor PP2, but is abrogated by actin polymerization inhibitor latrunculin A. We propose that Src kinase-independent formation of TCR microclusters in response to agonist MHC-peptide provides an actin-dependent scaffold for signal amplification.

Wound Healing on the Dorsal Hands: An Intrapatient Comparison of Primary Closure, Purse-String Closure, and Secondary Intention.

Nonmelanoma skin cancers are common on the dorsal hands where reserve tissue is limited. We highlight the case of an elderly man who had 3 nonmelanoma skin cancers on the left hand that were treated on the same day and left similar wounds. The wounds were repaired by primary closure, secondary intention, and purse-string circumferential closure. All wounds healed with excellent and essentially equivalent cosmetic results. For small shallow wounds on the dorsal hands, dermatologic surgeons should have confidence that secondary intention healing likely will lead to acceptable cosmetic and functional results.

Immunological synapse: a multi-protein signalling cellular apparatus for controlling gene expression.

The interaction of T cells with antigen-presenting cells is the hallmark of adaptive immunity. In vitro studies have described the formation of an immunological synapse between these cells, and intra-vital imaging has described in great detail the dynamics of these interactions. The immunological synapse has become a paradigm to study signals exchanged between the two cells. A wealth of information has been amassed regarding the localization of signalling molecules, their kinetics and the transcription factors they activate. We continue to discover mechanisms that cause receptors and signalling molecules to compartmentalize in the cell; however, the emerging challenge lies in understanding how the immunological synapse contributes to differentiation. Here, we review some of the transcription factors activated downstream of T-cell receptor signalling and discuss mechanisms by which antigen dose and affinity may influence differentiation. Antigen affinity might change the kind of transcription factors that are activated whereas antigen dose is likely to influence the temporal dynamics of the transcription factors. The immunological synapse is therefore likely to influence differentiation by modulating the trafficking of transcription factors and by promoting asymmetric cell division, an emerging concept.