Germline Variants in DNA Damage Repair Genes: An Emerging Role in the Era of Precision Medicine in Pancreatic Adenocarcinoma.

Pancreatic adenocarcinoma is a lethal disease that is projected to become the second most common cause of cancer deaths by 2030. The role of adjuvant therapy after surgical resection has been established by several clinical trials to prolong survival and improve outcomes. Multiagent chemotherapy seems to be the most promising approach to counteract early recurrence and improve survival; however, in the era of precision medicine, patient selection and individualized therapy seems to hold the key to desirable superior outcomes. Several cancer susceptibility genes have been proven to be associated with an increased risk of pancreatic cancer, both familial and sporadic cases. The role of genomic profiling for germline variants has been extensive and of limited clinical value, considering their low prevalence in pancreatic ductal adenocarcinoma (PDAC). However, an accumulating body of evidence from several studies in the past decade have successfully shown a recognizable value of germline variants in risk assessment and patient stratification. Recently, anti-PD-1 therapy (pembrolizumab) has been FDA-approved for use in solid malignancies with a Mismatch repair deficiency or high Microsatellite instability. Several trials have evaluated the role of poly (ADP-ribose) polymerase (PARP) inhibitors in patients harboring germline BRCA1/2 mutations. Finally, germline variants in DNA damage response genes and particularly deleterious ones have the potential to guide therapy after surgical resection and serve as biomarkers to predict survival. The dire need to address challenges for applying precision medicine in real-life clinical settings for PDAC patients lies in further characterizing the genetic and molecular processes through translational research.

Inter-rater agreement of rotator cuff tendon and muscle magnetic resonance imaging parameters evaluated preoperatively and during the first postoperative year following rotator cuff repair.

BACKGROUND: Magnetic resonance imaging (MRI) is standard of care for rotator cuff evaluation, with clinical interpretation usually limited to qualitative judgments. The reliability of MRI-based measurements and scoring systems has been evaluated only preoperatively or ≥6 months following rotator cuff repair, when repairs are in the later stages of healing. This study describes the MRI assessments and inter-rater agreement of various rotator cuff tendon and muscle parameters evaluated preoperatively and 4 times during the first postoperative year. METHODS: Two musculoskeletal radiologists independently assessed MRI scans of 42 patients preoperatively and 3, 12, 26, and 52 weeks after rotator cuff repair. Using standardized reading rules, readers assessed tendon integrity (5-point Sugaya classification), tear dimensions, muscle fat (5-point Goutallier classification) and atrophy (4-point Warner classification), muscle cross-sectional areas, and myotendinous junction distance. Raw exact agreement proportions, κ statistics, and correlation coefficients were used to quantify inter-rater agreement. RESULTS: Readers showed moderate to substantial above-chance agreement in scoring rotator cuff tendon integrity and supraspinatus muscle atrophy and good to excellent agreement on tear dimensions and muscle cross-sectional areas but only fair to moderate agreement for fatty infiltration and myotendinous junction distance. Only fatty infiltration grades evidenced observer bias. Inter-rater agreement did not appear time dependent. CONCLUSION: By use of defined reading rules in a research setting, MRI evaluations of rotator cuff tendon integrity, tear dimensions, muscle atrophy, and cross-sectional areas have reasonable reliability at all time points in the first postoperative year. However, the presence of clinically significant disagreements, even in such favorable circumstances, indicates the need for improved imaging tools for precise rotator cuff evaluation.

Variability of glenohumeral positioning and bone-to-tendon marker length measurements in repaired rotator cuffs from longitudinal computed tomographic imaging.

BACKGROUND: To address the need for more objective and quantitative measures of tendon healing in research studies, we intend to use computed tomography (CT) with implanted radiopaque markers on the repaired tendon to measure tendon retraction following rotator cuff repair. In our small prior study, retraction at 1-year follow-up averaged 16.1± 5.3 mm and exceeded 10.0 mm in 12 of 13 patients, and thus tendon retraction appears to be a common clinical phenomenon. This study's objectives were to assess, using 5 longitudinal CT scans obtained over 1 year following rotator cuff repair, the variability in glenohumeral positioning because of pragmatic variations in achieving perfect arm repositioning and to estimate the associated measurement variability in bone-to-tendon marker length measurements. METHODS: Forty-eight patients underwent rotator cuff repair with intraoperative placement of radiopaque tendon markers at the repair site. All patients had a CT scan with their arms at the side on the day of surgery and at 3, 12, 26, and 52 weeks postoperatively. Glenohumeral position (defined by the orientation and distance of the humerus with respect to the scapula) and bone-to-tendon marker lengths were measured from each scan. Within-patient variation in glenohumeral position measurements was described by their pooled within-patient standard deviations (SDs), and variation in bone-to-tendon marker lengths by their standard errors of measurement (SEMs) and 95% confidence level minimally detectable distances (MDD95) and changes (MDC95). RESULTS: The mean glenohumeral orientation from the 5 longitudinal CT scans averaged across the 48 patients was 12.6° abduction, 0.4° flexion, and -0.1° internal rotation. Within-patient SDs (95% confidence intervals) of glenohumeral orientation were 3.0° (2.7°-3.4°) in extension/flexion, 5.2° (4.6°-5.8°) in abduction/adduction, and 8.2° (7.3°-9.2°) in internal/external rotation. The SDs of glenohumeral distances were less than 1 mm in any direction. The estimated SEMs of bone-to-tendon lengths were consistent with a common value of 2.4 mm for any of the tendon markers placed across the repair, with MDD95 of 4.7 mm and MDC95 of 6.7 mm. CONCLUSION: Apparent tendon retraction of 5 mm or more, when measured as the distance from a tendon marker's day of surgery location to its new location on a volumetrically registered longitudinal CT scan, may be considered above the usual range of measurement variation. Tendon retraction measured using implanted radiopaque tendon markers offers an objective and sufficiently reliable means for quantifying the commonly expected changes in structural healing following rotator cuff repair.

Mechanistic insights into cancer cell killing through interaction of phosphodiesterase 3A and schlafen family member 12.

Cytotoxic molecules can kill cancer cells by disrupting critical cellular processes or by inducing novel activities. 6-(4-(Diethylamino)-3-nitrophenyl)-5-methyl-4,5-dihydropyridazin-3(2H)-one (DNMDP) is a small molecule that kills cancer cells by generation of novel activity. DNMDP induces complex formation between phosphodiesterase 3A (PDE3A) and schlafen family member 12 (SLFN12) and specifically kills cancer cells expressing elevated levels of these two proteins. Here, we examined the characteristics and covariates of the cancer cell response to DNMDP. On average, the sensitivity of human cancer cell lines to DNMDP is correlated with PDE3A expression levels. However, DNMDP could also bind the related protein, PDE3B, and PDE3B supported DNMDP sensitivity in the absence of PDE3A expression. Although inhibition of PDE3A catalytic activity did not account for DNMDP sensitivity, we found that expression of the catalytic domain of PDE3A in cancer cells lacking PDE3A is sufficient to confer sensitivity to DNMDP, and substitutions in the PDE3A active site abolish compound binding. Moreover, a genome-wide CRISPR screen identified the aryl hydrocarbon receptor-interacting protein (AIP), a co-chaperone protein, as required for response to DNMDP. We determined that AIP is also required for PDE3A-SLFN12 complex formation. Our results provide mechanistic insights into how DNMDP induces PDE3A-SLFN12 complex formation, thereby killing cancer cells with high levels of PDE3A and SLFN12 expression.

A novel radiopaque tissue marker for soft tissue localization and in vivo length and area measurements.

PURPOSE: The purpose of the study was to describe the characteristics and demonstrate proof-of-concept and clinical use of a barium sulfate infused polypropylene radiopaque tissue marker for soft tissue localization and in vivo measurement of lengths and areas. METHODS: Marker mechanical properties were evaluated by tensile tests. Biocompatibility was evaluated following 8-12 weeks' implantation in a pig model. Proof-of-concept of marker application was performed in a human cadaveric shoulder model, and methods for CT imaging and measurement of dimensions were established. Lastly, the method of clinical use of the markers was described in one patient undergoing arthroscopic rotator cuff repair (RCR). RESULTS: The radiopaque markers had a tensile strength of 28 ±4.7 N and were associated with minimal to mild inflammatory tissue reaction similar to polypropylene control. CT-based measurements showed relatively high precisions for lengths (0.66 mm), areas (6.97 mm2), and humeral orientation angles (2.1°) in the cadaveric model, and demonstrated 19 ±3 mm medio-lateral tendon retraction and 227 ±3 mm2 increase in tendon area in the patient during 26 weeks following RCR. No radiographic leaching, calcification or local adverse events were observed. CONCLUSIONS: The radiopaque tissue marker was biocompatible and had adequate strength for handling and affixation to soft tissues using standard suturing techniques. The marker could be used with low-dose, sequential CT imaging to quantitatively measure rotator cuff tendon retractions with clinically acceptable accuracy. We envision the radiopaque tissue marker to be useful for soft tissue localization and in vivo measurement of tissue and organ dimensions following surgery.

Development of an Arthroscopic Joint Capsule Injury Model in the Canine Shoulder.

BACKGROUND: The natural history of rotator cuff tears can be unfavorable as patients develop fatty infiltration and muscle atrophy that is often associated with a loss of muscle strength and shoulder function. To facilitate study of possible biologic mechanisms involved in early degenerative changes to rotator cuff muscle and tendon tissues, the objective of this study was to develop a joint capsule injury model in the canine shoulder using arthroscopy. METHODS: Arthroscopic surgical methods for performing a posterior joint capsulectomy in the canine shoulder were first defined in cadavers. Subsequently, one canine subject underwent bilateral shoulder joint capsulectomy using arthroscopy, arthroscopic surveillance at 2, 4 and 8 weeks, and gross and histologic examination of the joint at 10 weeks. RESULTS: The canine subject was weight-bearing within eight hours after index and follow-up surgeries and had no significant soft tissue swelling of the shoulder girdle or gross lameness. Chronic synovitis and macroscopic and microscopic evidence of pathologic changes to the rotator cuff bony insertions, tendons, myotendinous junctions and muscles were observed. CONCLUSIONS: This study demonstrates feasibility and proof-of-concept for a joint capsule injury model in the canine shoulder. Future work is needed to define the observed pathologic changes and their role in the progression of rotator cuff disease. Ultimately, better understanding of the biologic mechanisms of early progression of rotator cuff disease may lead to clinical interventions to halt or slow this process and avoid the more advanced and often irreversible conditions of large tendon tears with muscle fatty atrophy.

Does augmentation with a reinforced fascia patch improve rotator cuff repair outcomes?

BACKGROUND: Scaffold devices are used to augment rotator cuff repairs in humans. While the strength of a novel poly-L-lactic acid-reinforced (human) fascia patch has been documented, it is unclear whether such patches will enhance the strength or likelihood of healing of rotator cuff repairs. QUESTIONS/PURPOSES: In a canine shoulder model, we asked: Do tendon repairs augmented with a reinforced fascia patch have (1) increased biomechanical properties at Time 0 and (2) less tendon retraction and increased cross-sectional area and biomechanical properties after 12 weeks of healing compared to repairs without augmentation? (3) Do the biomechanical properties of tendon repairs reach normal values by 12 weeks of healing? And (4) is the host response associated with use of the reinforced fascia patch biocompatible? METHODS: Eleven dogs underwent bilateral shoulder surgery with partial release and acute repair of the infraspinatus tendon, one shoulder with augmentation and one without augmentation. Repair retraction, cross-sectional area, biomechanical properties, and biocompatibility were assessed at 12 weeks. RESULTS: At Time 0, the mean ± SD ultimate load of augmented repairs was 296 ± 130 N (46% ± 25%) more than nonaugmented repairs, with no difference in stiffness between groups. At 12 weeks, the ultimate load of augmented repairs averaged 192 ± 213 N (15% ± 16%) less than nonaugmented repairs, with no difference in stiffness between groups. At the tendon repair site at 12 weeks, the fascia patch showed a biocompatible host tissue response. CONCLUSIONS: The biomechanical properties of repairs augmented with a reinforced fascia patch demonstrated greater ultimate load at Time 0 than nonaugmented repairs but remained essentially unchanged after 12 weeks of healing, despite improvements in the ultimate load of nonaugmented controls in the same time frame.

Estimation of dynamic, in vivo soft-tissue deformation: experimental technique and application in a canine model of tendon injury and repair.

Outcomes after rotator cuff surgery are typically assessed with measures of strength, joint motion, or pain, but these measures do not provide a direct assessment of tissue function as healing progresses. To address this limitation, this manuscript describes biplane X-ray analysis as a technique for quantifying in vivo soft-tissue deformation. Tantalum beads were implanted in the humerus and infraspinatus tendon in a canine model of tendon injury and repair. Biplane X-ray images were acquired during treadmill trotting and tissue deformation was estimated from the three-dimensional bead positions. Changes over time were characterized by the mean, range, and normalized range (i.e., range/mean) of interbead distance. Intact tendon repair tissue demonstrated significant decreases over time in the mean (p = 0.003), range (p = 0.001), and normalized range (p = 0.001) of interbead distance. Failed tendon repair tissue demonstrated significant decreases over time in the range (p = 0.05) and normalized range (p = 0.04) of interbead distance. In an uninjured control, differences over time in the interbead distance parameters were not detected. This approach is a promising technique for estimating changes over time in soft-tissue deformation. These preliminary data indicate appreciable differences between normal tendons, intact repairs, and failed repairs.

Rotator cuff repair augmentation in a canine model with use of a woven poly-L-lactide device.

BACKGROUND: Despite advances in surgical treatment options, failure rates of rotator cuff repair have continued to range from 20% to 90%. Hence, there is a need for new repair strategies that provide effective mechanical reinforcement of rotator cuff repair as well as stimulate and enhance the intrinsic healing potential of the patient. The purpose of this study was to evaluate the extent to which augmentation of acute repair of rotator cuff tendons with a newly designed poly-L-lactide repair device would improve functional and biomechanical outcomes in a canine model. METHODS: Eight adult, male mongrel dogs (25 to 30 kg) underwent bilateral shoulder surgery. One shoulder underwent tendon release and repair only, and the other was subjected to release and repair followed by augmentation with the repair device. At twelve weeks, tendon retraction, cross-sectional area, stiffness, and ultimate load of the repair site were measured. Augmented repairs underwent histologic assessment of biocompatibility. In addition, eight pairs of canine cadaver shoulders underwent infraspinatus injury and repair with and without device augmentation with use of identical surgical procedures and served as time-zero biomechanical controls. Eight unpaired, canine cadaver shoulders were included as normal biomechanical controls. RESULTS: At time zero, repair augmentation significantly increased the ultimate load (23%) (p = 0.034) but not the stiffness of the canine infraspinatus tendon repair. At twelve weeks, the poly-L-lactide scaffold was observed to be histologically biocompatible, and augmented repairs demonstrated significantly less tendon retraction (p = 0.008) and significantly greater cross-sectional area (137%), stiffness (26%), and ultimate load (35%) than did repairs that had not been augmented (p < 0.001, p = 0.002, and p = 0.009, respectively). CONCLUSIONS: While limiting but not eliminating tendon repair retraction, the augmentation device provided a tendon-bone bridge and scaffold for host tissue deposition and ingrowth, resulting in improved biomechanical function of the repair at twelve weeks.

Assessment of the canine model of rotator cuff injury and repair.

Animal shoulder models are used to systematically investigate the factors influencing rotator cuff injury and repair. Each model has advantages and disadvantages that must be considered in the context of the specific research questions being asked. This study evaluated the utility of the canine model for studies of acute, full-thickness rotator cuff tendon injury and repair. We found that time-zero failure load is dependent on the suture type and configuration used for repair. Acute, full-width tendon repairs fail anatomically within the first days after surgery in the canine model, regardless of suture type, suture configuration, or postoperative protocol. Robust scar tissue forms in the gap between the failed tendon end and the humerus, which can be visually, mechanically, and histologically misconstrued as tendon if an objective test of repair connectivity is not performed. We conclude that a full-width injury and repair model in the canine will provide a rigorous test of whether a new repair strategy or postoperative protocol, such as casting or temporary muscle paralysis, can maintain repair integrity in a high-load environment. Alternatively, a partial-width tendon injury model allows loads to be shared between the tendon repair and the remaining intact portion of the infraspinatus tendon and prohibits complete tendon retraction. Thus a partial-width injury in the canine may model the mechanical environment of many single tendon tears in the human injury condition and warrants further investigation.

Commercial extracellular matrix scaffolds for rotator cuff tendon repair. Biomechanical, biochemical, and cellular properties.

BACKGROUND: We are not aware of any in vitro study comparing the biomechanical, biochemical, and cellular properties of commercial extracellular matrix materials marketed for rotator cuff tendon repair. In this study, the properties of GraftJacket, TissueMend, Restore, and CuffPatch were quantified and compared with each other. The elastic moduli were also compared with that of normal canine infraspinatus tendon. METHODS: Samples were tested from different manufacturing lots of four materials: GraftJacket (ten lots), TissueMend (six), Restore (ten), and CuffPatch (six). The Kruskal-Wallis test was used to compare thickness, stiffness, and modulus as well as hydroxyproline, chondroitin/dermatan sulfate glycosaminoglycan, hyaluronan, and DNA contents among these matrices. The moduli of the extracellular matrices were also compared with those of normal canine infraspinatus tendon. RESULTS: All four extracellular matrices required 10% to 30% stretch before they began to carry substantial load. Their maximum moduli were realized in their linear region at 30% to 80% strain. The elastic moduli of all four commercial matrices were an order of magnitude lower than that of canine infraspinatus tendon. TissueMend had significantly higher DNA content than the other three matrices (p<0.0001), although both Restore and GraftJacket also had measurable amounts of DNA. CONCLUSIONS: Our data demonstrate chemical and mechanical differences among the four commercial extracellular matrices that we evaluated. Probably, the source (dermis or small intestine submucosa), species (human, porcine, or bovine), age of the donor (fetal or adult), and processing of these matrices all contribute to the unique biophysical properties of the delivered product. The biochemical composition of commercial extracellular matrices is similar to that of tendon. However, the elastic moduli of these materials are an order of magnitude lower than that of tendon, suggesting a limited mechanical role in augmentation of tendon repair.