Enhancing the Efficacy of HIPEC Through Bromelain: A Preclinical Investigation in Appendiceal Cancer.

INTRODUCTION: Appendiceal cancer (AC) excessive mucin production is a barrier to heated intraperitoneal chemotherapy (HIPEC) drug delivery. Bromelain is a pineapple stem extract with mucolytic properties. We explored bromelain treatment effects against mucinous AC in a patient-derived tumor organoid (PTO) model and an AC cell line. PATIENTS AND METHODS: PTOs were fabricated from tumor specimens obtained from patients with AC undergoing cytoreductive surgery with HIPEC. PTOs underwent HIPEC treatment with bromelain, cisplatin, and mitomycin C (MMC) at 37 °C and 42 °C with and without bromelain pretreatment. RESULTS: From October 2020 to May 2023, 16 specimens were collected from 13 patients with low-grade (12/16, 75%) and high-grade AC (4/16, 25%). The mucin-depleting effects of bromelain were most significant in combination with N-acetylcysteine (NAC) compared with bromelain (47% versus 10%, p = 0.0009) or NAC alone (47% versus 12.8%, p = 0.0027). Bromelain demonstrated > 31% organoid viability reduction at 60 min (p < 0.001) and > 66% in 48 h (p < 0.0001). Pretreatment with bromelain increased cytotoxicity of both cisplatin and MMC HIPEC conditions by 31.6% (p = 0.0001) and 35.5% (p = 0.0001), respectively. Ki67, CK20, and MUC2 expression decreased after bromelain treatment; while increased caspase 3/7 activity and decreased Bcl-2 (p = 0.009) and Bcl-xL (p = 0.01) suggest induction of apoptosis pathways. Furthermore, autophagy proteins LC3A/B I (p < 0.03) and II (p < 0.031) were increased; while ATG7 (p < 0.01), ATG 12 (p < 0.04), and Becline 1(p < 0.03), expression decreased in bromelain-treated PTOs. CONCLUSIONS: Bromelain demonstrates cytotoxicity and mucolytic activity against appendiceal cancer organoids. As a pretreatment agent, it potentiates the cytotoxicity of multiple HIPEC regimens, potentially mediated through programmed cell death and autophagy.

Long-Term Survival in Patients Treated with Cytoreduction and Heated Intraperitoneal Chemotherapy for Peritoneal Mesothelioma at a Single High-Volume Center.

BACKGROUND: Malignant peritoneal mesothelioma (MPM) is a rare diagnosis with a dismal prognosis if untreated. Cytoreductive surgery with hyperthermic intraperitoneal chemotherapy (CRS-HIPEC) is shown to significantly improve survival. Our institution is uniquely positioned to report long-term outcomes in MPM with CRS-HIPEC, due to our robust peritoneal surface disease program existing over the past three decades. METHODS: Our prospectively maintained, single-institution database of CRS-HIPEC cases was reviewed, identifying 111 consecutive patients with MPM over 28 years (1993-2021). Prognostic, operative, and pathologic factors were reviewed. Overall survival (OS) and conditional survival (CS) analyses were performed. RESULTS: The average age was 55.1 years; 58.6% of patients were male; 17 of 111 patients (15.3%) had a second CRS-HIPEC. At first CRS-HIPEC, the average PCI score was 18.7, and the perfusate drugs were platinum-based (72.1%) and mitomycin C (27.9%). The resection status at first CRS-HIPEC was R2a (46.4%), followed by R0-1 (29.1%), and R2b-c (24.5%). Median OS was 3.3 years for the entire cohort, with 75th and 25th percentiles at 10.7 months and 10.6 years. Median CS was improved if patients survived to the 1-year postoperative mark (4.9 years, p < 0.01) and trended toward further improvement with each passing year. If 3-year postoperative survival was achieved, the median CS improved to 6.1 years. CONCLUSIONS: This represents one of the largest and lengthiest, single-center, longitudinal, case series of peritoneal mesothelioma treated with CRS-HIPEC. The OS suggests efficacy for CRS-HIPEC for MPM. Long-term survival improves significantly after patients achieve the 1-year, postoperative mark.

National Evaluation of HIV Service Resource Allocation in Tanzania.

Using time-driven activity-based costing (TDABC), we examined resource allocation and costs for HIV services throughout Tanzania at patient and facility levels. This national, cross-sectional analysis of 22 health facilities quantified costs and resources associated with 886 patients receiving care for five HIV services: antiretroviral therapy, prevention of mother-to-child transmission, HIV testing and counseling, voluntary medical male circumcision, and pre-exposure prophylaxis. We also documented total provider-patient interaction time, the cost of services with and without inclusion of consumables, and conducted fixed-effects multivariable regression analyses to examine patient- and facility-level correlates of costs and provider-patient time. Findings showed that resources and costs for HIV care varied significantly throughout Tanzania, including as a function of patient- and facility-level characteristics. While some variation may be preferable (e.g., needier patients received more resources), other areas suggested a lack of equity (e.g., wealthier patients received more provider time) and presented opportunities to optimize care delivery protocols.

Unit cost repositories for health program planning and evaluation: a report on research in practice with lessons learned.

BACKGROUND: Most low- and middle-income countries have limited access to cost data that meets the needs of health policy-makers and researchers in health intervention areas including HIV, tuberculosis, and immunization. Unit cost repositories (UCRs)-searchable databases that systematically codify evidence from costing studies-have been developed to reduce the effort required to access and use existing costing information. These repositories serve as public resources and standard references, which can improve the consistency and quality of resource needs projections used for strategic planning and resource mobilization. UCRs also enable analysis of cost determinants and more informed imputation of missing cost data. This report examines our experiences developing and using seven UCRs (two global, five country-level) for cost projection and research purposes. DISCUSSION: We identify advances, challenges, enablers, and lessons learned that might inform future work related to UCRs. Our lessons learned include: (1) UCRs do not replace the need for costing expertise; (2) tradeoffs are required between the degree of data complexity and the useability of the UCR; (3) streamlining data extraction makes populating the UCR with new data easier; (4) immediate reporting and planning needs often drive stakeholder interest in cost data; (5) developing and maintaining UCRs requires dedicated staff time; (6) matching decision-maker needs with appropriate cost data can be challenging; (7) UCRs must have data quality control systems; (8) data in UCRs can become obsolete; and (9) there is often a time lag between the identification of a cost and its inclusion in UCRs. CONCLUSIONS: UCRs have the potential to be a valuable public good if kept up-to-date with active quality control and adequate support available to end-users. Global UCR collaboration networks and greater control by local stakeholders over global UCRs may increase active, sustained use of global repositories and yield higher quality results for strategic planning and resource mobilization.

A document analysis of the South African national guidelines for medical male circumcision and the implications for infant and child male circumcision decision-making.

South Africa's 2016 medical male circumcision (MMC) guidelines ("the guidelines") provide direction for the MMC programme's implementation in South Africa. The aim of our document analysis was to assess the guidelines, particularly in lieu of changing guidance from WHO and PEPFAR. We then assessed how the guidelines might be applied to infant and child male circumcision (ICMC). The analysis was performed by reviewing South Africa's guidelines, along with international documents used in developing those guidelines, to identify the historical development of the guidelines, as well as the implications for MMC and ICMC decision-making within the South African context. The following principles were analysed within the context of South Africa's guidelines: (1) quality and safety; (2) informed consent; (3) confidentiality; (4) human rights; and (5) accessibility of services. Tthe document analysis also identified ambiguities that exist in the guidelines, particularly regarding consent, recognising religious or cultural exemptions, and guaranteeing the best interests of the child. South Africa's MMC guidelines could benefit from incorporating common definitions to assist with interpretation and understanding, thus preventing confusion and controversy among programme planners, parents and boys. The guidelines were made available in 2016 and recommendations for circumcision have evolved as new research and information has become available. South Africa's National Department of Health should review and update these guidelines, with a focus on both MMC and ICMC issues, so that they reflect the most up-to-date, accurate information available, to avoid inconsistent practices, risks, and litigation in the management of the programme. This study was situated within a qualitative paradigm and applied a social choice theory perspective to make sense of the MMC guidelines. The study concludes that future policy revisions should develop a broader understanding of the complex medical male circumcision decision-making process, particularly faced by parents.

Patient-Specific Sarcoma Organoids for Personalized Translational Research: Unification of the Operating Room with Rare Cancer Research and Clinical Implications.

INTRODUCTION: Sarcoma clinical outcomes have been stagnant for decades due to heterogeneity of primaries, lack of comprehensive preclinical models, and rarity of disease. We hypothesized that engineering hydrogel-based sarcoma organoids directly from the patient without xenogeneic extracellular matrices (ECMs) or growth factors is routinely feasible and allows rare tumors to remain viable as avatars for personalized research. METHODS: Surgically resected sarcomas (angiosarcomas, leiomyosarcoma, gastrointestinal stromal tumor, liposarcoma, myxofibrosarcoma, dermatofibrosarcoma protuberans [DFSP], and pleiomorphic abdominal sarcoma) were dissociated and incorporated into a hyaluronic acid and collagen-based ECM hydrogel and screened for chemotherapy efficacy. A subset of organoids was enriched with a patient-matched immune system for screening of immunotherapy efficacy (iPTOs). Response to treatment was assessed using LIVE/DEAD staining and metabolic assays. RESULTS: Sixteen sarcomas were biofabricated into three-dimensional (3D) patient-specific sarcoma organoids with a 100% success rate. Average time from organoid development to initiation of drug testing was 7 days. Enrichment of organoids with immune system components derived from either peripheral blood mononuclear cells or lymph node cells was performed in 10/16 (62.5%) patients; 4/12 (33%) organoids did not respond to chemotherapy, while response to immunotherapy was observed in 2/10 (20%) iPTOs. CONCLUSIONS: A large subset of sarcoma organoids does not exhibit response to chemotherapy or immunotherapy, as currently seen in clinical practice. Routine development of sarcoma hydrogel-based organoids directly from the operating room is a feasible platform, allowing for such rare tumors to remain viable for personalized translational research.

Estimating the cost and efficiency gain of rolling out a multi-month dispensing programme for antiretroviral treatment in Tanzania.

Background: Globally, efforts to curtail the HIV pandemic are growing. The Joint United Nations Programme on HIV and AIDS (UNAIDS) and partners set the 95-95-95 targets to be achieved by 2025. Tanzania's ongoing transition from single-month ARV to longer multi-month dispensing (MMD) involves significant planning and shifts in existing resources, including health commodities, clinical staff and storage space. This study aimed at evaluating the costs and efficiency gains of rolling out MMD compared to the prior monthly dispending (MD) standard of care before the new guidelines.Methods: The analysis employed a health provider perspective utilising prior costing data collected to estimate cost of treatment for HIV/AIDS, including salaries, laboratory costs, antiretroviral drugs, other supplies and overhead costs. The projections were run from 2018 to 2030 using the Spectrum package for Tanzania.Results: Our model estimated that total treatment cost without MMD (including salaries, laboratory costs, antiretroviral drugs, other supplies, and overhead costs) is estimated to rise from USD 189 million in 2018 to USD 244 million in 2030. The introduction of a six-month MMD would lead to the total annual facility-based treatment costs being reduced to USD 205 million in 2030. When comparing MD to a six-month MMD, the total savings over the 13-year period would be USD 425 million. The introduction of six-month MMD for stable patients would reduce the average cost from USD 180 to USD 156 per patient per year if stable patients were only required to make six-monthly visit.Conclusions: The introduction of differentiated service delivery models (DSDMs) and MMD is already contributing to significant cost savings for Tanzania and will continue to do so as the country puts more stable patients on MMD. The potential gains from MMD implantation could further be harnessed if retention of treatment and viral suppression monitoring are prioritised.

Personalized Identification of Optimal HIPEC Perfusion Protocol in Patient-Derived Tumor Organoid Platform.

BACKGROUND: Chemotherapy dosing duration and perfusion temperature vary significantly in HIPEC protocols. This study investigates patient-derived tumor organoids as a platform to identify the most efficacious perfusion protocol in a personalized approach. PATIENTS AND METHODS: Peritoneal tumor tissue from 15 appendiceal and 8 colon cancer patients who underwent CRS/HIPEC were used for personalized organoid development. Organoids were perfused in parallel at 37 and 42 °C with low- and high-dose oxaliplatin (200 mg/m2 over 2 h vs. 460 mg/m2 over 30 min) and MMC (40 mg/3L over 2 h). Viability assays were performed and pooled for statistical analysis. RESULTS: An adequate organoid number was generated for 75% (6/8) of colon and 73% (11/15) of appendiceal patients. All 42 °C treatments displayed lower viability than 37 °C treatments. On pooled analysis, MMC and 200 mg/m2 oxaliplatin displayed no treatment difference for either appendiceal or colon organoids (19% vs. 25%, p = 0.22 and 27% vs. 31%, p = 0.55, respectively), whereas heated MMC was superior to 460 mg/m2 oxaliplatin in both primaries (19% vs. 54%, p < 0.001 and 27% vs. 53%, p = 0.002, respectively). In both appendiceal and colon tumor organoids, heated 200 mg/m2 oxaliplatin displayed increased cytotoxicity as compared with 460 mg/m2 oxaliplatin (25% vs. 54%, p < 0.001 and 31% vs. 53%, p = 0.008, respectively). CONCLUSIONS: Organoids treated with MMC or 200 mg/m2 heated oxaliplatin for 2 h displayed increased susceptibility in comparison with 30-min 460 mg/m2 oxaliplatin. Optimal perfusion protocol varies among patients, and organoid technology may offer a platform for tailoring HIPEC conditions to the individual patient level.

Model of Patient-Specific Immune-Enhanced Organoids for Immunotherapy Screening: Feasibility Study.

INTRODUCTION: We hypothesized that engineering a combined lymph node/melanoma organoid from the same patient would allow tumor, stroma, and immune system to remain viable for personalized immunotherapy screening. METHODS: Surgically obtained matched melanoma and lymph node biospecimens from the same patient were transferred to the laboratory and washed with saline, antibiotic, and red blood cell lysis buffer. Biospecimens were dissociated, incorporated into an extracellular matrix (ECM)-based hydrogel system, and biofabricated into three dimensional (3D) mixed melanoma/node organoids. Cells were not sorted, so as to preserve tumor heterogeneity, including stroma and immune cell components, resulting in immune-enhanced patient tumor organoids (iPTOs). Organoid sets were screened in parallel with nivolumab, pembrolizumab, ipilimumab, and dabrafenib/trametinib for 72 h. LIVE/DEAD staining and quantitative metabolism assays recorded relative drug efficacy. Histology and immunohistochemistry were used to compare tumor melanoma cells with organoid melanoma cells. Lastly, node-enhanced iPTOs were employed to activate patient-matched peripheral blood T cells for killing of tumor cells in naïve PTOs. RESULTS: Ten biospecimen sets obtained from eight stage III and IV melanoma patients were reconstructed as symbiotic immune/tumor organoids between September 2017 and June 2018. Successful establishment of viable organoid sets was 90% (9/10), although organoid yield varied with biospecimen size. Average time from organoid development to initiation of immunotherapy testing was 7 days. In three patients for whom a node was not available, it was substituted with peripheral blood mononuclear cells. iPTO response to immunotherapy was similar to specimen clinical response in 85% (6/7) patients. In an additional pilot study, peripheral T cells were circulated through iPTOs, and subsequently transferred to naïve PTOs from the same patient, resulting in tumor killing, suggesting a possible role of iPTOs in generating adaptive immunity. CONCLUSION: Development of 3D mixed immune-enhanced tumor/node organoids is a feasible platform, allowing individual patient immune system and tumor cells to remain viable for studying of personalized immunotherapy response.

Appendiceal Cancer Patient-Specific Tumor Organoid Model for Predicting Chemotherapy Efficacy Prior to Initiation of Treatment: A Feasibility Study.

INTRODUCTION: We have hypothesized that biofabrication of appendiceal tumor organoids allows for a more personalized clinical approach and facilitates research in a rare disease. METHODS: Appendiceal cancer specimens obtained during cytoreduction with hyperthermic intraperitoneal chemotherapy procedures (CRS/HIPEC) were dissociated and incorporated into an extracellular matrix-based hydrogel system as three-dimensional (3D), patient-specific tumor organoids. Cells were not sorted, preserving tumor heterogeneity, including stroma and immune cell components. Following establishment of organoid sets, chemotherapy drugs were screened in parallel. Live/dead staining and quantitative metabolism assays recorded which chemotherapies were most effective in killing cancer cells for a specific patient. Maintenance of cancer phenotypes were confirmed by using immunohistochemistry. RESULTS: Biospecimens from 12 patients were applied for organoid development between November 2016 and May 2018. Successful establishment rate of viable organoid sets was 75% (9/12). Average time from organoid development to chemotherapy testing was 7 days. These tumors included three high-grade appendiceal (HGA) and nine low-grade appendiceal (LGA) primaries obtained from sites of peritoneal metastasis. All tumor organoids were tested with chemotherapeutic agents exhibited responses that were either similar to the patient response or within the variability of the expected clinical response. More specifically, HGA tumor organoids derived from different patients demonstrated variable chemotherapy tumor-killing responses, whereas LGA organoids tested with the same regimens showed no response to chemotherapy. One LGA set of organoids was immune-enhanced with cells from a patient-matched lymph node to demonstrate feasibility of a symbiotic 3D reconstruction of a patient matched tumor and immune system component. CONCLUSIONS: Development of 3D appendiceal tumor organoids is feasible even in low cellularity LGA tumors, allowing for individual patient tumors to remain viable for research and personalized drug screening.

HIV treatment support services in Tanzania: a cost and efficiency analysis at facility and community levels.

Differentiated service delivery models provide an opportunity to reduce the cost of HIV treatment while increasing the focus on advanced patients. Also, stable patients who need less attention can visit facilities less frequently, saving time and money and reducing overcrowding. Tanzania needs treatment support services that can improve the quality of life, ensure adherence and result in viral suppression. Treatment support services can be provided through a variety of models, including at the community level, through the facility or through some combination of the two. Understanding the cost of each model is essential for policy-makers who must allocate resources. Data from neighbouring countries suggests that community-level support services can be beneficial to patients, especially in reducing loss to follow-up and death. Though community-based HIV services are available in Tanzania, uncertainty about the costs of these models remain. This study assessed the costs of treatment support services at 27 sites in Tanzania. The cost analysis found that the average unit cost of treatment support services is US$39 per patient per year. The analysis found that community-based models have clear advantages with regard to the number of patients that can be reached with support services. Costing data indicated that community-based models are also less expensive. The lack of data on retention limited any conclusions about whether community-based models are the most effective within the Tanzanian context.

Meta-analysis of average costs of HIV testing and counselling and voluntary medical male circumcision across thirteen countries.

Objective: Explore facility-level average costs per client of HIV testing and counselling (HTC) and voluntary medical male circumcision (VMMC) services in 13 countries.Methods: Through a literature search we identified studies that reported facility-level costs of HTC or VMMC programmes. We requested the primary data from authors and standardised the disparate data sources to make them comparable. We then conducted descriptive statistics and a meta-analysis to assess the cost variation among facilities. All costs were converted to 2017 US dollars ($).Results: We gathered data from 14 studies across 13 countries and 772 facilities (552 HTC, 220 VMMC). The weighted average unit cost per client served was $15 (95% CI 12, 18) for HTC and $59 (95% CI 45, 74) for VMMC. On average, 38% of the mean unit cost for HTC corresponded to recurrent costs, 56% to personnel costs, and 6% to capital costs. For VMMC, 41% of the average unit cost corresponded to recurrent costs, 55% to personnel costs, and 4% to capital costs. We observed unit cost variation within and between countries, and lower costs in higher scale categories in all interventions.

Cisplatin induces mitochondrial deficits in Drosophila larval segmental nerve.

Cisplatin is an effective chemotherapy drug that induces peripheral neuropathy in cancer patients. In rodent dorsal root ganglion neurons, cisplatin binds nuclear and mitochondrial DNA (mtDNA) inducing DNA damage and apoptosis. Platinum-mtDNA adducts inhibit mtDNA replication and transcription leading to mitochondrial degradation. Cisplatin also induces climbing deficiencies associated with neuronal apoptosis in adult Drosophila melanogaster. Here we used Drosophila larvae that express green fluorescent protein in the mitochondria of motor neurons to observe the effects of cisplatin on mitochondrial dynamics and function. Larvae treated with 10µg/ml cisplatin had normal survival with deficiencies in righting and heat sensing behavior. Behavior was abrogated by, the pan caspase inhibitor, p35. However, active caspase 3 was not detected by immunostaining. There was a 27% decrease in mitochondrial membrane potential and a 42% increase in reactive oxygen species (ROS) in mitochondria along the axon. Examination of mitochondrial axonal trafficking showed no changes in velocity, flux or mitochondrial length. However, cisplatin treatment resulted in a greater number of stationary organelles caused by extended pausing during axonal motility. These results demonstrate that cisplatin induces behavior deficiencies in Drosophila larvae, decreased mitochondrial activity, increased ROS production and mitochondrial pausing without killing the larvae. Thus, we identified particular aspects of mitochondrial dynamics and function that are affected in cisplatin-induced peripheral neuropathy and may represent key therapeutic targets.

A reductionist metastasis-on-a-chip platform for in vitro tumor progression modeling and drug screening.

Current animal and 2-D cell culture models employed in metastasis research and drug discovery remain poor mimics of human cancer physiology. Here we describe a "metastasis-on-a-chip" system allowing real time tracking of fluorescent colon cancer cells migrating from hydrogel-fabricated gut constructs to downstream liver constructs within a circulatory fluidic device system that responds to environmental manipulation and drug treatment. Devices consist of two chambers in which gut and liver constructs are housed independently, but are connected in series via circulating fluid flow. Constructs were biofabricated with a hyaluronic acid-based hydrogel system, capable of a variety of customizations, inside of which representative host tissue cells were suspended and metastatic colon carcinoma tumor foci were created. The host tissue of the constructs expressed normal epithelial markers, which the tumor foci failed to express. Instead, tumor regions lost membrane-bound adhesion markers, and expressed mesenchymal and proliferative markers, suggesting a metastatic phenotype. Metastatic tumor foci grew in size, eventually disseminating from the intestine construct and entering circulation, subsequently reaching in the liver construct, thus mimicking some of the migratory events observed during metastasis. Lastly, we demonstrated the ability to manipulate the system, including chemically modulating the hydrogel system mechanical properties and administering chemotherapeutic agents, and evaluated the effects of these parameters on invasive tumor migration. These results describe the capability of this early stage metastasis-on-a-chip system to model several important characteristics of human metastasis, thereby demonstrating the potential of the platform for making meaningful advances in cancer investigation and drug discovery. Biotechnol. Bioeng. 2016;113: 2020-2032. © 2016 Wiley Periodicals, Inc.