Effect of palliative radiation dose on symptom response in metastatic sarcomas.

Purpose: Palliative radiotherapy (RT) plays a crucial role in alleviating symptoms associated with metastatic sarcoma. However, there is a lack of consensus on the optimal palliative radiation dose and fractionation for metastatic sarcomas. We analyzed the association between biologically effective radiation dose and symptom response for patients who underwent palliative RT for metastatic sarcomas. Methods and materials: We retrospectively identified patients with metastatic sarcoma treated with palliative RT between 1999 and 2021 at our institution. We assessed the association between equivalent dose in 2 Gy fractions (EQD2) with an α/ß of three and symptom relief or overall survival (OS) using univariable and multivariable analyses. Results: Of the 198 metastatic sites treated, the most common indications for palliative radiation were pain (n = 181, 91 %) and compression of adjacent structures (n = 16, 8 %). In our analysis, an EQD2 of > 20 Gy was associated with greater rates of short-term symptom relief (n = 143, 85 %) at the RT site compared to an EQD2 of ≤ 20 Gy (n = 14, 54 %, P = 0.001) with no reports of grade 3 or higher toxicity. However, there was no significant improvement in short-term symptom relief for higher radiation doses. Patients treated with an EQD2 of ≤ 20 Gy had a significantly worse performance status, but there was no significant difference in overall survival based on EQD2 on multivariable analysis. Conclusions: An EQD2 ≤ 20 Gy (e.g., 8 Gy in 1 fraction) provided inadequate palliative benefit in this series. An EQD2 > 20 Gy resulted in greater rates of symptom palliation in metastatic sarcomas, but further dose escalation did not improve symptom response or durability. These findings suggest standard palliative regimens such as 20 Gy in 5 fractions (EQD2 of 28 Gy) are effective for patients with metastatic sarcomas.

Chest wall pain after single-fraction thoracic stereotactic ablative Radiotherapy: Dosimetric analysis from the iSABR trial.

BACKGROUND AND PURPOSE: Concerns over chest wall toxicity has led to debates on treating tumors adjacent to the chest wall with single-fraction stereotactic ablative radiotherapy (SABR). We performed a secondary analysis of patients treated on the prospective iSABR trial to determine the incidence and grade of chest wall pain and modeled dose-response to guide radiation planning and estimate risk. MATERIALS AND METHODS: This analysis included 99 tumors in 92 patients that were treated with 25 Gy in one fraction on the iSABR trial which individualized dose by tumor size and location. Toxicity events were prospectively collected and graded based on the CTCAE version 4. Dose-response modeling was performed using a logistic model with maximum likelihood method utilized for parameter fitting. RESULTS: There were 22 grade 1 or higher chest wall pain events, including five grade 2 events and zero grade 3 or higher events. The volume receiving at least 11 Gy (V11Gy) and the minimum dose to the hottest 2 cc (D2cc) were most highly correlated with toxicity. When dichotomized by an estimated incidence of ≥ 20 % toxicity, the D2cc > 17 Gy (36.6 % vs. 3.7 %, p < 0.01) and V11Gy > 28 cc (40.0 % vs. 8.1 %, p < 0.01) constraints were predictive of chest wall pain, including among a subset of patients with tumors abutting or adjacent to the chest wall. CONCLUSION: For small, peripheral tumors, single-fraction SABR is associated with modest rates of low-grade chest wall pain. Proximity to the chest wall may not contraindicate single fractionation when using highly conformal, image-guided techniques with sharp dose gradients.

Standards for fecal microbiota transplant: Tools and therapeutic advances.

Fecal microbiota transplantation (FMT) has been demonstrated to be efficacious in preventing recurrent Clostridioides difficile (C. difficile) infections, and is being investigated for treatment of several other diseases including inflammatory bowel disease, cancer, obesity, liver disease, and diabetes. To speed up the translation of FMT into clinical practice as a safe and standardized therapeutic intervention, additional evidence-based technical and regulatory guidance is needed. To this end in May of 2022, the International Alliance for Biological Standardization (IABS) and the BIOASTER Microbiology Technology Institute hosted a second webinar to discuss key issues still impeding the advancement and standardization of FMT. The goal of this two-day webinar was to provide a forum for scientific experts to share and discuss data and key challenges with one another. Discussion included a focus on the evaluation of safety, efficacy, clinical trial design, reproducibility and accuracy in obtained microbiome measurements and data reporting, and the potential for standardization across these areas. It also focused on increasing the application potential and visibility of FMT beyond treating C. difficile infections.

Less than whole uterus irradiation for patients with locally advanced cervical cancer.

INTRODUCTION: Current consensus guidelines for definitive cervical cancer intensity modulated radiation therapy (IMRT) recommend inclusion of the entire uterus within the clinical target volume, however this is debated. We aimed to evaluate outcomes of patients with cervical cancer who were treated with less than whole uterus irradiation. METHODS: We identified 109 patients with FIGO Stage IB-IVA cervical cancer treated definitively with concurrent chemoradiation, including IMRT and brachytherapy, from 2010 to 2022 at a single institution where the practice was to include the gross cervix tumor with an internal target volume with differences in bladder filing accounted for, plus additional 5 mm planning target volume (PTV) margin. Local, regional, and distant recurrences were analyzed using competing risk methods, and a Wilcoxon rank sum test was performed to assess differences in dose to organs at risk based on the proportion of the uterus included in the PTV, with the median proportion of the uterus included (75 %) used as the cut-point. RESULTS: The median follow-up time was 65 months (range 3-352 months). The 2-year cumulative incidence of LR for the entire cohort was 4.2 % (95 % confidence interval [CI] 1.3-9.7). Compared with patients who had ≥ 75 % of the uterus included in the PTV, patients who had < 75 % of the uterus included in the PTV had significantly lower bowel D200cc (p = 0.02). The cumulative incidence of local failure (LR) was not significantly different between the two groups. CONCLUSIONS: Including less than the whole uterus for definitive cervix cancer IMRT does not seem to compromise local control. Less than whole uterus irradiation could be considered for carefully selected cervix cancer patients to decrease bowel dose and possible treatment-related toxicity.

Impact of lung transplantation on diastolic dysfunction in recipients with pretransplant pulmonary hypertension.

OBJECTIVE: Pulmonary hypertension can cause left ventricular diastolic dysfunction through ventricular interdependence. Moreover, diastolic dysfunction has been linked to adverse outcomes after lung transplant. The impact of lung transplant on diastolic dysfunction in recipients with pretransplant pulmonary hypertension is not defined. In this cohort, we aimed to assess the prevalence of diastolic dysfunction, the change in diastolic dysfunction after lung transplant, and the impact of diastolic dysfunction on lung transplant outcomes. METHODS: In a large, single-center database from January 2011 to September 2021, single or bilateral lung transplant recipients with pulmonary hypertension (mean pulmonary artery pressure > 20 mm Hg) were retrospectively identified. Those without a pre- or post-transplant echocardiogram within 1 year were excluded. Diastolic dysfunction was diagnosed and graded according to the American Society of Echocardiography 2016 guideline on assessment of diastolic dysfunction (present, absent, indeterminate). McNemar's test was used to examine association between diastolic dysfunction pre- and post-transplant. Kaplan-Meier and Cox regression analysis were used to assess associations between pre-lung transplant diastolic dysfunction and post-lung transplant 1-year outcomes, including mortality, major adverse cardiac events, and bronchiolitis obliterans syndrome grade 1 or higher-free survival. RESULTS: Of 476 primary lung transplant recipients, 205 with pulmonary hypertension formed the study cohort (mean age, 56.6 ± 11.9 years, men 61.5%, mean pulmonary artery pressure 30.5 ± 9.8 mm Hg, left ventricular ejection fraction < 55% 9 [4.3%]). Pretransplant, diastolic dysfunction was present in 93 patients (45.4%) (grade I = 8, II = 84, III = 1), absent in 16 patients (7.8%), and indeterminate in 89 patients (43.4%), and 7 patients (3.4%) had missing data. Post-transplant, diastolic dysfunction was present in 7 patients (3.4%) (grade I = 2, II = 5, III = 0), absent in 164 patients (80.0%), and indeterminate in 15 patients (7.3%), and 19 patients (9.3%) had missing data. For those with diastolic dysfunction grades in both time periods (n = 180), there was a significant decrease in diastolic dysfunction post-transplant (148/169 patients with resolved diastolic dysfunction; McNemar's test P < .001). Pretransplant diastolic dysfunction was not associated with major adverse cardiac events (hazard ratio [HR], 1.08, 95% CI, 0.72-1.62; P = .71), bronchiolitis obliterans syndrome-free survival (HR, 0.67, 95% CI, 0.39-1.56; P = .15), or mortality (HR, 0.70, 95% CI, 0.33-1.46; P = .34) at 1 year. CONCLUSIONS: Diastolic dysfunction is highly prevalent in lung transplant candidates with normal left ventricular systolic function and pulmonary hypertension, and resolves in most patients after lung transplant regardless of patient characteristics. Pre-lung transplant diastolic dysfunction was not associated with adverse lung or cardiac outcomes after lung transplant. Collectively, these findings suggest that the presence of diastolic dysfunction in lung transplant recipients with pulmonary hypertension has no prognostic significance, and as such diastolic dysfunction and the associated clinical syndrome of heart failure with preserved ejection fraction should not be considered a relative contraindication to lung transplant in such patients.

Characteristics and Outcomes of De Novo Genitourinary Malignancy in Solid Organ Transplant Recipients at the University of Minnesota.

BACKGROUND: Studies examining outcomes of genitourinary malignancy (GU) in the solid organ transplant (SOT) population predominantly focus on renal transplant recipients and consist of relatively small cohorts. We aimed to expand knowledge of the characteristics and outcomes of de novo GU malignancies in all patients with SOT at a large tertiary center. METHODS: The SOT database was queried for recipients with de novo bladder, renal cell, and prostate malignancy, and a retrospective chart review was performed. Descriptive statistics and Kaplan-Meier survival estimates were calculated. Cox proportional hazards regression was used for multivariate modeling of predictive factors in the development of GU malignancy. RESULTS: Solid organ transplant recipients with de novo bladder malignancy comprised 64.3% with high grade and 38.1% with advanced stage (≥T2) disease at initial diagnosis. Only 3.7% of patients with de novo renal cell carcinoma presented with metastatic disease, and 13.6% with localized disease developed recurrences. The most common stage in de novo prostate cancer patients was pT3 (52.2%). Kaplan-Meier estimates (95% CI) for 5-year overall (OS) and cancer-specific survival (CSS) were 44.12% (31.13-62.52) and 80.80% (68.85-94.81) for bladder, 78.90% (68.93-90.30) and 96.61% (92.10-100.00) for renal cell, and 81.18% (72.01-91.51) and 96.16% (90.95-100.00) for prostate cancer, respectively. Age at transplant and time from transplant to cancer diagnosis were predictive of de novo bladder cancer OS (P = .042 and .021, respectively). CONCLUSION: To our knowledge, this is the largest single-center cohort examined for GU malignancy after SOT. Bladder and renal cell cancer had worse OS but similar CSS as historical rates for nontransplant patients. De novo prostate cancer had similar CSS.

Structural Basis of the Allosteric Inhibition of Human ABCG2 by Nanobodies.

ABCG2 is an ATP-binding cassette transporter that exports a wide range of xenobiotic compounds and has been recognized as a contributing factor for multidrug resistance in cancer cells. Substrate and inhibitor interactions with ABCG2 have been extensively studied and small molecule inhibitors have been developed that prevent the export of anticancer drugs from tumor cells. Here, we explore the potential for inhibitors that target sites other than the substrate binding pocket of ABCG2. We developed novel nanobodies against ABCG2 and used functional analyses to select three inhibitory nanobodies (Nb8, Nb17 and Nb96) for structural studies by single particle cryo-electron microscopy. Our results showed that these nanobodies allosterically bind to different regions of the nucleotide binding domains. Two copies of Nb8 bind to the apex of the NBDs preventing them from fully closing. Nb17 binds near the two-fold axis of the transporter and interacts with both NBDs. Nb96 binds to the side of the NBD and immobilizes a region connected to key motifs involved in ATP binding and hydrolysis. All three nanobodies prevent the transporter from undergoing conformational changes required for substrate transport. These findings advance our understanding of the molecular basis of modulation of ABCG2 by external binders, which may contribute to the development of a new generation of inhibitors. Furthermore, this is the first example of modulation of human multidrug resistance transporters by nanobodies.

Recipient and kidney graft outcomes of deceased donors with human immunodeficiency virus in the United States.

BACKGROUND: The HIV Organ Policy Equity (HOPE) act afforded transplantation of organs from donors who have HIV. Herein we compared the long-term outcomes of recipients with HIV by donor HIV testing status. METHODS: Using the Scientific Registry of Transplant Recipients, we identified all primary adult kidney transplant recipients who were HIV-positive between 1/1/16-12/31/21. Recipients were grouped into three cohorts according to the donor HIV status based on antibody (Ab) and nucleic acid testing (NAT): Donor Ab-/NAT- (n = 810), Donor Ab+ /NAT- (n = 98), and Donor Ab+/NAT+ (n = 90). We compared recipient and death-censored graft survival (DCGS) by donor HIV testing status using Kaplan-Meier curves and Cox proportional hazards regression, censored at 3 years posttransplant. Secondary outcomes were delayed graft function (DGF) and the following 1-year outcomes: acute rejection, re-hospitalization, and serum creatinine. RESULTS: In Kaplan-Meier analyses, patient survival and DCGS were similar by donor HIV status (log rank p = .667; log rank p = .388). DGF occurred more frequently in donors with HIV Ab-/NAT- testing compared with Ab+/NAT- or Ab+/NAT+ testing (38.0% vs. 28.6% vs. 26.7%, p = .028). Average dialysis time before transplant was twice as long for recipients who received organs from donors with Ab-/NAT- testing (p < .001). Acute rejection, re-hospitalization and serum creatinine at 12 months did not differ between the groups. CONCLUSIONS: Patient and allograft survival for recipients living with HIV remains comparable irrespective of donor HIV testing status. Utilizing kidneys from deceased donors with HIV Ab+/NAT- or Ab+/NAT+ testing shortens dialysis time prior to transplant.

BK DNAemia in pediatric kidney transplant recipients: Predictors and outcomes.

BACKGROUND: Pediatric data on risk factors and the clinical course of BK DNAemia are limited. We aimed to determine the effects of BK DNAemia on transplant outcomes and delineate the safety and efficacy of various treatment approaches. METHODS: This retrospective-cohort study included 161 transplants (age ≤ 21 years) performed at a single center between 1/1/2012 and 1/1/2020. We used Cox proportional models to evaluate the effects of BK DNAemia on patient survival (PS), graft survival (GS), and acute rejection (AR), using BK as a time-dependent covariate. We also assessed the effects of pharmacological intervention on BK DNAemia duration using intervention as a time-dependent covariate. RESULTS: BK-free survival was 69.1% at 1-year and 54.6% at 3-year posttransplant. After multivariate adjustment, BK DNAemia was associated with young age at transplant (aHR, age 5-<12 vs. ≥12 (years): 2.5 (1.4-4.5); p = .001) and steroid-based immunosuppression (IS) (aHR: 2.2 [1.1-4.5]; p = .03). We found no effect of DNAemia on AR (aHR: 1.25; p = .5), PS (aHR: 2.85; p = .22), and GS (aHR: 0.56; p = .41). Of 70 patients with DNAemia, 22 (31.4%) received no treatment, 20 (28.6%) received IS reduction alone, and 28 patients (40%) received treatment with at least one pharmacological agent (leflunomide, IVIG, ciprofloxacin, cidofovir). Sixty-three patients (90%) cleared DNAemia with median time to resolution of 2.4 months (IQR:1.4-5.6). We found no significant effect of BK-directed pharmacological treatment on time to resolution (aHR: 0.64;p = .13). BK-directed agents were well tolerated. CONCLUSIONS: BK DNAemia is associated with a young age at transplant and steroid-based maintenance IS. We found no effect of BK DNAemia on AR, GS, and PS.

Orchialgia After Living Donor Nephrectomy: An Underreported Entity.

Laparoscopic donor nephrectomy (LDN) offers advantages to the donor. The reported incidence of testicular pain after LDN varies in the literature ranging from 3% to 55%. Methods: A survey was sent to 322 male LDN patients who donated from February 5, 2009, to February 5, 2019. The survey assessed if the donor had testicular pain or saw an additional medical professional after donation. Results: Of the 322 surveyed, 147 (46%) responses were received. Of those who had a left nephrectomy, 39% had testicular pain; 23.8% of those patients had testicular swelling in addition. Of those who had pain, laterality of kidney donated did not impact if the patient had pain, pain onset, pain level, or pain duration. Of those who donated their right kidney, 35% had testicular pain, and 16.7% of those patients reported testicular swelling in addition. Twenty-seven symptomatic patients sought additional medical care for the testicular symptoms postdonation. Seven (25%) had hydroceles, 2 (7%) had testicular cysts, 1 had a urinary tract infection, and 16 (59%) had reassurance or no additional procedures provided. Conclusions: Our results suggest that orchialgia is not as uncommon as previously thought and may be one of the most common minor complications experienced by male donors.

Preemptive Second Kidney Transplant Outcomes by Induction Type in the United States.

BACKGROUND: The role of induction in preemptive second kidney recipients is unclear. We examined the association between induction therapy and the long-term graft and recipient survival in the settings of tacrolimus and mycophenolate maintenance. METHODS: We identified all preemptive second kidney transplant recipients between 2000 and 2020 in the Scientific Registry of Transplant Recipients. We excluded those with missing or mixed induction regimens and positive crossmatch. We grouped recipients by induction type into 3 groups: anti-thymocyte globulin (n = 1442), alemtuzumab (n = 362), and interleukin-2 receptor antagonist (IL-2RA; n = 481). We generated Kaplan-Meier curves of the recipient and death-censored graft survival (DCGS) with follow-up censored at 10 years. We used multivariable Cox proportional hazards models to examine the association between induction and the above outcomes. We adjusted the models for recipient and donor variables. RESULTS: Rates of delayed graft function, rejection, hospitalization, and post-transplant lymphoproliferative disorder at one year were not statistically different. Recipient survival did not vary by induction type in the Kaplan-Meier analysis (log-rank P = .189) or in the multivariable model. However, DCGS was the lowest in the Alemtuzumab group (log-rank P = .01). In the multivariable models, alemtuzumab was associated with a 57% increased risk of graft loss (1.57, 95% confidence interval (1.08, 2.30), P = .019) compared to anti-thymocyte. Live-donor kidneys were associated with significantly better recipient survival and DCGS. CONCLUSIONS: Compared to anti-thymocyte induction, alemtuzumab, but not IL-2RA, was associated with inferior graft survival in preemptive second transplant recipients discharged on tacrolimus and mycophenolate.

Phylogenomics of the genus Glycine sheds light on polyploid evolution and life-strategy transition.

Polyploidy and life-strategy transitions between annuality and perenniality often occur in flowering plants. However, the evolutionary propensities of polyploids and the genetic bases of such transitions remain elusive. We assembled chromosome-level genomes of representative perennial species across the genus Glycine including five diploids and a young allopolyploid, and constructed a Glycine super-pangenome framework by integrating 26 annual soybean genomes. These perennial diploids exhibit greater genome stability and possess fewer centromere repeats than the annuals. Biased subgenomic fractionation occurred in the allopolyploid, primarily by accumulation of small deletions in gene clusters through illegitimate recombination, which was associated with pre-existing local subgenomic differentiation. Two genes annotated to modulate vegetative-reproductive phase transition and lateral shoot outgrowth were postulated as candidates underlying the perenniality-annuality transition. Our study provides insights into polyploid genome evolution and lays a foundation for unleashing genetic potential from the perennial gene pool for soybean improvement.

TAR30, a homolog of the canonical plant TTTAGGG telomeric repeat, is enriched in the proximal chromosome regions of peanut (Arachis hypogaea L.).

Telomeres are the physical ends of eukaryotic linear chromosomes that play critical roles in cell division, chromosome maintenance, and genome stability. In many plants, telomeres are comprised of TTTAGGG tandem repeat that is widely found in plants. We refer to this repeat as canonical plant telomeric repeat (CPTR). Peanut (Arachis hypogaea L.) is a spontaneously formed allotetraploid and an important food and oil crop worldwide. In this study, we analyzed the peanut genome sequences and identified a new type of tandem repeat with 10-bp basic motif TTTT(C/T)TAGGG named TAndem Repeat (TAR) 30. TAR30 showed significant sequence identity to TTTAGGG repeat in 112 plant genomes suggesting that TAR30 is a homolog of CPTR. It also is nearly identical to the telomeric tandem repeat in Cestrum elegans. Fluorescence in situ hybridization (FISH) analysis revealed interstitial locations of TAR30 in peanut chromosomes but we did not detect visible signals in the terminal ends of chromosomes as expected for telomeric repeats. Interestingly, different TAR30 hybridization patterns were found between the newly induced allotetraploid ValSten and its diploid wild progenitors. The canonical telomeric repeat TTTAGGG is also present in the peanut genomes and some of these repeats are closely adjacent to TAR30 from both cultivated peanut and its wild relatives. Overall, our work identifies a new homolog of CPTR and reveals the unique distributions of TAR30 in cultivated peanuts and wild species. Our results provide new insights into the evolution of tandem repeats during peanut polyploidization and domestication.

Lung Transplant Recipients With Prior Coronary Artery Bypass Grafting Are at Increased Risk of Death and Early Perioperative Hemorrhage.

Prior coronary artery bypass grafting (CABG) has been considered a relative contraindication to lung transplantation due to the atherosclerotic disease burden and technical challenges. We hypothesized that lung transplant recipients with prior CABG have increased mortality compared to recipients without prior CABG. Further, the causes of death are different for lung transplant recipients with prior CABG vs without CABG. The Scientific Registry of Transplant Recipients database was queried to define the survival and causes of death of lung transplant recipients with or without CABG during the Lung Allocation Score era from May 5, 2005 to December 31, 2015. The primary end-points were all-cause mortality at 1 year and 5 years, as well as mortality due to major causes of death. This retrospective study cohort included a total of 13,064 lung transplant recipients, of whom 319 patients had previously undergone CABG, representing 2.4% of all transplants. Patients without prior CABG were more likely to have undergone bilateral lung transplantation compared to those with prior CABG (61.2 % vs 15.7%, P < 0.001). Among patients with prior CABG, single right lung transplant was most common. Overall patient survival at 1 year was 76.8% for lung transplant recipients with prior CABG and 85.4% for patients without prior CABG. Freedom from death due to graft failure at 1 and 5 years in patients with a prior CABG was 93.1% and 76.2% respectively, cardiac and/or cerebrovascular disease 96.2% and 88.5% respectively, and hemorrhage 97.9% and 97.5% respectively. In a multivariate Cox regression model utilizing time-dependent coefficients for recipient age, prior CABG, among several other risk factors, was associated with increased mortality within 1 year. Prior CABG is associated with short- and long-term mortality in lung transplant recipients with history of CABG despite the majority of these patients undergoing single lung transplantation vs bilateral lung transplantation. Graft failure and/or pulmonary causes are the most common cause of death regardless of whether or not the lung transplant recipient had prior CABG, but patients with prior CABG are at increased risk of death due to graft failure, cardiac or cerebrovascular disease, and hemorrhage.

Structures of ABCG2 under turnover conditions reveal a key step in the drug transport mechanism.

ABCG2 is a multidrug transporter that affects drug pharmacokinetics and contributes to multidrug resistance of cancer cells. In previously reported structures, the reaction cycle was halted by the absence of substrates or ATP, mutation of catalytic residues, or the presence of small-molecule inhibitors or inhibitory antibodies. Here we present cryo-EM structures of ABCG2 under turnover conditions containing either the endogenous substrate estrone-3-sulfate or the exogenous substrate topotecan. We find two distinct conformational states in which both the transport substrates and ATP are bound. Whereas the state turnover-1 features more widely separated NBDs and an accessible substrate cavity between the TMDs, turnover-2 features semi-closed NBDs and an almost fully occluded substrate cavity. Substrate size appears to control which turnover state is mainly populated. The conformational changes between turnover-1 and turnover-2 states reveal how ATP binding is linked to the closing of the cytoplasmic side of the TMDs. The transition from turnover-1 to turnover-2 is the likely bottleneck or rate-limiting step of the reaction cycle, where the discrimination of substrates and inhibitors occurs.

Structural Basis of Drug Recognition by the Multidrug Transporter ABCG2.

ABCG2 is an ATP-binding cassette (ABC) transporter whose function affects the pharmacokinetics of drugs and contributes to multidrug resistance of cancer cells. While its interaction with the endogenous substrate estrone-3-sulfate (E1S) has been elucidated at a structural level, the recognition and recruitment of exogenous compounds is not understood at sufficiently high resolution. Here we present three cryo-EM structures of nanodisc-reconstituted, human ABCG2 bound to anticancer drugs tariquidar, topotecan and mitoxantrone. To enable structural insight at high resolution, we used Fab fragments of the ABCG2-specific monoclonal antibody 5D3, which binds to the external side of the transporter but does not interfere with drug-induced stimulation of ATPase activity. We observed that the binding pocket of ABCG2 can accommodate a single tariquidar molecule in a C-shaped conformation, similar to one of the two tariquidar molecules bound to ABCB1, where tariquidar acts as an inhibitor. We also found single copies of topotecan and mitoxantrone bound between key phenylalanine residues. Mutagenesis experiments confirmed the functional importance of two residues in the binding pocket, F439 and N436. Using 3D variability analyses, we found a correlation between substrate binding and reduced dynamics of the nucleotide binding domains (NBDs), suggesting a structural explanation for drug-induced ATPase stimulation. Our findings provide additional insight into how ABCG2 differentiates between inhibitors and substrates and may guide a rational design of new modulators and substrates.

Evolution of Homeologous Gene Expression in Polyploid Wheat.

Polyploidization has played a prominent role in the evolutionary history of plants. Two recent and sequential allopolyploidization events have resulted in the formation of wheat species with different ploidies, and which provide a model to study the effects of polyploidization on the evolution of gene expression. In this study, we identified differentially expressed genes (DEGs) between four BBAA tetraploid wheats of three different ploidy backgrounds. DEGs were found to be unevenly distributed among functional categories and duplication modes. We observed more DEGs in the extracted tetraploid wheat (ETW) than in natural tetraploid wheats (TD and TTR13) as compared to a synthetic tetraploid (AT2). Furthermore, DEGs showed higher Ka/Ks ratios than those that did not show expression changes (non-DEGs) between genotypes, indicating DEGs and non-DEGs experienced different selection pressures. For A-B homeolog pairs with DEGs, most of them had only one differentially expressed copy, however, when both copies of a homeolog pair were DEGs, the A and B copies were more likely to be regulated to the same direction. Our results suggest that both cis- and inter-subgenome trans-regulatory changes are important drivers in the evolution of homeologous gene expression in polyploid wheat, with ploidy playing a significant role in the process.

Recurrence of Hepatocellular Carcinoma in Hepatitis C Virus (HCV) Liver Transplant Recipients Treated with Pretransplant Direct-Acting Antiviral (DAA) Therapy.

BACKGROUND: Direct-acting antivirals (DAAs) have revolutionized the treatment of hepatitis C virus (HCV). The impact of DAAs on recurrence of hepatocellular carcinoma (HCC) after liver transplantation (LT) remains uncertain. OBJECTIVE: We aimed to evaluate the risk of HCC recurrence in LT recipients cleared of HCV with DAAs at the time of LT compared to a control group of LT recipients who were viremic at the time of LT. METHODS: The study was a single-center, retrospective cohort study of patients undergoing LT for HCV-related HCC from 2002 to 2017. We compared time to post-LT HCC recurrence in patients with a sustained virological response (SVR) from DAAs prior to LT (DAA group) to patients who were viremic at LT (HCV+ group) using Kaplan-Meier analysis. We performed a secondary analysis comparing post-LT HCC recurrence in the DAA group to LT recipients with SVR from interferon-based treatment prior to LT (IFN group). RESULTS: One hundred fifty-one patients underwent LT for HCC related to HCV: 34 patients in DAA group, 95 patients in HCV+ group, and 22 in IFN group. Kaplan-Meier estimates of being HCC free were 96.2, 96.2, and 78.8% at 6, 12, and 24 months in DAA group, respectively, and 100, 98.6, and 95.8% at 6, 12, and 24 months in the HCV+ group, respectively; p = 0.08. There was no difference observed for HCC recurrence between the DAA and IFN groups. In a multivariate Cox proportional hazards model, DAA use increased the risk of post-LT HCC recurrence (HR 5.2, 95% CI 0.9-29.81, p = 0.07). CONCLUSIONS: A strong trend was observed on both Kaplan-Meier and multivariate analyses toward increased post-LT HCC recurrence in patients who achieved SVR prior to LT with DAAs compared to patients who were viremic at LT. Caution is required when considering pre-LT treatment of HCV with DAAs in patients with HCC.

Risk of Genitourinary Malignancy in the Renal Transplant Patient.

OBJECTIVE: To better understand the risk of genitourinary malignancies in the renal transplant patient. Currently, no consensus exists regarding screening and intervention, with much of the clinical decision-making based on historical practices established before recent progress in immunosuppression protocols and in genitourinary cancer diagnosis and management. METHODS: A database of all solid organ transplants performed at the University of Minnesota from 1984 to 2019 was queried for renal transplant recipients in whom development of subsequent urologic malignancies (prostate, bladder, renal, penile, and testicular cancer) was found. RESULTS: Among 6172 renal transplant recipients examined, cumulative incidence of all cancers of genitourinary etiology are presented over an average follow-up time of 10 years. Kidney cancer (combined graft and native), prostate cancer, and bladder cancer each demonstrated respective 30-year incidence of 4.6%, 8.7%, and 1.5% from the time of transplant. By comparison, age-matched data from the Surveillance, Epidemiology, and End Results database demonstrated 30-year cumulative incidence of 1.1%, 11.1%, and 1.7% for kidney cancer, prostate cancer, and bladder cancer respectively. The predominant genitourinary cancer was renal cell cancer, both of the native and of the transplanted kidney (native, n = 64; transplanted, n =11), followed by prostate cancer (n = 63), and bladder cancer (n = 37). CONCLUSION: In this closely followed cohort of renal transplant recipients, renal cancer occurs at a higher incidence rate than in the non-transplanted population, while a lower rate of prostate cancer was found, with bladder cancer demonstrating a comparable cumulative incidence between transplant patients and the national age-matched population.

Separate Effect of Perioperative Recombinant Human Factor VIIa Administration and Packed Red Blood Cell Transfusions on Midterm Survival in Lung Transplantation Recipients.

OBJECTIVE: The purpose of this study was to determine the relationship between blood product transfusion, with or without recombinant human activated factor VIIa, and survival after lung transplantation. DESIGN: Retrospective analysis of a single center with follow-up out to 6 years post-transplantation. SETTING: Single-center academic lung transplantation program. PARTICIPANTS: The study comprised 265 adult patients who underwent single or bilateral sequential lung transplantation from March 2011 to June 2017. INTERVENTIONS: Overall survival using Kaplan-Meier curves was compared among the following 3 cohorts: those not transfused with blood products, those transfused with blood products, and those given blood products and recombinant human activated factor VIIa. Cox proportional hazards regression was used to estimate hazard ratios (HRs), confidence intervals (CIs), and p values. MEASUREMENTS AND MAIN RESULTS: Seventy-eight patients received no packed red blood cell transfusions, 149 received packed red blood cell transfusions, and 38 received both packed red blood cell transfusions and recombinant human activated factor VII. Packed red blood cell transfusion was associated with an increased risk of mortality that did not reach statistical significance (HR 2.168, CI 0.978-4.805; p = 0.057). Additional packed red blood cells beyond 15 U were associated with worsened survival (HR 1.363, CI 1.137-1.633; p = 0.001), but recombinant human activated factor VIIa did not increase the risk of mortality. CONCLUSION: Blood product transfusion during and after lung transplantation is associated with decreased survival, especially with large-volume transfusions. Survival is not worse with recombinant human activated factor VIIa administration, but additional studies are needed to determine whether recombinant human activated factor VIIa administration reduces the need for blood product transfusions.