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Objective: We integrate a new approach to chemosensitivity data for clinically-relevant regimen matching, and demonstrate the relationship with clinical outcomes in a large PDO biobank., Summary Background Data: Pancreatic ductal adenocarcinoma (PDAC) usually recurs following potentially curative resection. Prior studies related patient-derived organoid (PDO) chemosensitivity with clinical responses., Methods: PDOs were established from pre-treatment biopsies in a multi-institution clinical trial (n=21) and clinical specimens at a high-volume pancreatectomy center (n=74, of which 48 were pre-treated). PDO in vitro chemosensitivities to standard-of-care chemotherapeutics (pharmacotypes) were matched to potential clinically-relevant regimens by a weighted nearest-neighbors analysis. Clinical outcomes were then compared for patients who had well-matched versus poorly-matched treatment according to this metric., Results: Our function matched 91% of PDOs to a standard-of-care regimen (9% pan-resistant). PDOs poorly-matched to the neoadjuvant regimen received would have matched to an alternative in 34% of cases. Patients receiving neoadjuvant chemotherapy well-matched to their pharmacotype experienced improved CA 19-9 response (60% decreased to normal when well-matched, 29% when poorly-matched, P<0.05) and lymph node down-staging (33% N0 after poorly-matched, 69% after well-matched, P<0.05). Patients receiving both well-matched neoadjuvant and adjuvant chemotherapy experienced improved recurrence-free- and overall survival (median RFS 8.5 mo poorly-matched, 15.9 mo well-matched, P<0.05; median OS 19.5 vs. 30.3 mo, P<0.05)., Conclusion: In vitro PDO pharmacotyping can inform PDAC therapy selection. We demonstrate improved outcomes including survival for patients treated with regimens well-matched to their PDO chemosensitivities. A subsequent prospective study using PDO pharmacotype matching could improve oncologic outcomes and improve quality of life by avoiding therapies not expected to be effective., Competing Interests: The below disclosures are outside the context of the submitted work: TTS is the CEO and co-owner of Healthfund Finland and reports an interview honorarium from Boeringer Ingelheim. E.M.J is a paid consultant for Adaptive Biotech, Achilles, DragonFly, Candel Therapeutics, Genocea, and Roche. She receives funding from Lustgarten Foundation and Bristol Myer Squibb. She is the Chief Medical Advisor for Lustgarten and SAB advisor to the Parker Institute for Cancer Immunotherapy (PICI) and for the C3 Cancer Institute. She is a founding member of Abmeta. E.J.F is on the SAB for Resistance Biology, Consultant for Mestag Therapeutics and Merck. J.W.Z. reports grant funding from Genentech outside the submitted work. All other authors declare no potential conflict of interest., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Background: The aim of the authors was to determine whether socioeconomic vulnerability is associated with community water fluoridation (CWF)., Methods: The authors used US Census Bureau data to create 4 county-level vulnerability markers (percentages non-White, Hispanic or Latino, below the federal poverty threshold, education below high school), obtained county-level CWF data from the Washington State Department of Health, and evaluated associations using Spearman rank correlation coefficient and the Kruskal-Wallis rank sum test. The authors then interviewed 122 community members in Washington (December 2022-March 2023) and analyzed the interview data inductively., Results: A higher percentage of non-White people at the county level was associated with a significantly higher level of CWF (Spearman rank correlation coefficient, 0.55; 95% CI, 0.29 to 0.82; P < .001), whereas county-level poverty was associated with significantly lower CWF (Spearman rank correlation coefficient, -0.36; 95% CI, -0.70 to -0.03; P = .02). High school completion was not associated with county-level CWF. Significantly larger proportions of Hispanics and Latinos lived in counties with higher CWF (P < .05). From the interviews, more participants thought tap water was healthy than unhealthy, but 41% had mixed feelings. Similarly, more participants thought CWF was acceptable than unacceptable, with 35% reporting mixed feelings. Negative views about tap water and CWF were more common among non-White participants., Conclusions: People in racially and ethnically diverse communities in Washington appear to have greater access to CWF, whereas those in lower-income communities have poorer access., Practical Implications: CWF is an important population-level strategy to prevent caries. Additional work is needed to improve access to CWF, especially for people from low-income communities., Competing Interests: Disclosures None of the authors reported any disclosures., (Copyright © 2024 American Dental Association. Published by Elsevier Inc. All rights reserved.)

Costimulation blockade (CoB)-based immunotherapy is a promising alternative to immunosuppression for transplant recipients; however, the current limited understanding of the factors that impact its efficacy restrains its clinical applicability. In this context, pro- and anti-inflammatory cytokines are being recognized as having an impact on T cell activation beyond effector differentiation. This study aims at elucidating the impact of direct IL-10 signaling in T cells on CoB outcomes. We used a full-mismatch skin transplantation model where recipients had a T cell-restricted expression of a dominant negative IL-10 receptor (10R-DN), alongside anti-CD154 as CoB therapy. Unlike wild-type recipients, 10R-DN mice failed to benefit from CoB. This accelerated graft rejection correlated with increased accumulation of T cells producing TNF-α, IFN-γ, and IL-17. In vitro experiments indicated that while lack of IL-10 signaling did not change the ability of anti-CD154 to modulate alloreactive T cell proliferation, the absence of this pathway heightened T H 1 effector cell differentiation. Furthermore, deficiency of IL-10 signaling in T cells impaired Treg induction, a hallmark of anti-CD154 therapy. Overall, these findings unveil an important and novel role of IL-10 signaling in T cells that defines the success of CoB therapies and identifies a target pathway for obtaining robust immunoregulation., Competing Interests: Disclosure The authors of this manuscript have no conflicts of interest to disclose as described by the American Journal of Transplantation.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy characterized by an immunosuppressive tumor microenvironment enriched with cancer-associated fibroblasts (CAF). This study used a convergence approach to identify tumor cell and CAF interactions through the integration of single-cell data from human tumors with human organoid coculture experiments. Analysis of a comprehensive atlas of PDAC single-cell RNA sequencing data indicated that CAF density is associated with increased inflammation and epithelial-mesenchymal transition (EMT) in epithelial cells. Transfer learning using transcriptional data from patient-derived organoid and CAF cocultures provided in silico validation of CAF induction of inflammatory and EMT epithelial cell states. Further experimental validation in cocultures demonstrated integrin beta 1 (ITGB1) and vascular endothelial factor A (VEGFA) interactions with neuropilin-1 mediating CAF-epithelial cell cross-talk. Together, this study introduces transfer learning from human single-cell data to organoid coculture analyses for experimental validation of discoveries of cell-cell cross-talk and identifies fibroblast-mediated regulation of EMT and inflammation., Significance: Adaptation of transfer learning to relate human single-cell RNA sequencing data to organoid-CAF cocultures facilitates discovery of human pancreatic cancer intercellular interactions and uncovers cross-talk between CAFs and tumor cells through VEGFA and ITGB1., (©2024 American Association for Cancer Research.)

Purpose: Targeting solid tumors with chimeric antigen receptor (CAR) T cells remains challenging due to heterogenous target antigen expression, antigen escape, and the immunosuppressive tumor microenvironment (TME). Pancreatic cancer is characterized by a thick stroma generated by cancer-associated fibroblasts (CAF), which may contribute to the limited efficacy of mesothelin-directed CAR T cells in early-phase clinical trials. To provide a more favorable TME for CAR T cells to target pancreatic ductal adenocarcinoma (PDAC), we generated T cells with an antimesothelin CAR and a secreted T-cell-engaging molecule (TEAM) that targets CAF through fibroblast activation protein (FAP) and engages T cells through CD3 (termed mesoFAP CAR-TEAM cells)., Experimental Design: Using a suite of in vitro, in vivo, and ex vivo patient-derived models containing cancer cells and CAF, we examined the ability of mesoFAP CAR-TEAM cells to target PDAC cells and CAF within the TME. We developed and used patient-derived ex vivo models, including patient-derived organoids with patient-matched CAF and patient-derived organotypic tumor spheroids., Results: We demonstrated specific and significant binding of the TEAM to its respective antigens (CD3 and FAP) when released from mesothelin-targeting CAR T cells, leading to T-cell activation and cytotoxicity of the target cell. MesoFAP CAR-TEAM cells were superior in eliminating PDAC and CAF compared with T cells engineered to target either antigen alone in our ex vivo patient-derived models and in mouse models of PDAC with primary or metastatic liver tumors., Conclusions: CAR-TEAM cells enable modification of tumor stroma, leading to increased elimination of PDAC tumors. This approach represents a promising treatment option for pancreatic cancer., (©2024 American Association for Cancer Research.)

Pancreatic cancer is more prevalent in older individuals and often carries a poorer prognosis for them. The relationship between the microenvironment and pancreatic cancer is multifactorial, and age-related changes in nonmalignant cells in the tumor microenvironment may play a key role in promoting cancer aggressiveness. Because fibroblasts have profound impacts on pancreatic cancer progression, we investigated whether age-related changes in pancreatic fibroblasts influence cancer growth and metastasis. Proteomics analysis revealed that aged fibroblasts secrete different factors than young fibroblasts, including increased growth/differentiation factor 15 (GDF-15). Treating young mice with GDF-15 enhanced tumor growth, whereas aged GDF-15 knockout mice showed reduced tumor growth. GDF-15 activated AKT, rendering tumors sensitive to AKT inhibition in an aged but not young microenvironment. These data provide evidence for how aging alters pancreatic fibroblasts and promotes tumor progression, providing potential therapeutic targets and avenues for studying pancreatic cancer while accounting for the effects of aging., Significance: Aged pancreatic fibroblasts secrete GDF-15 and activate AKT signaling to promote pancreatic cancer growth, highlighting the critical role of aging-mediated changes in the pancreatic cancer microenvironment in driving tumor progression. See related commentary by Isaacson et al., p. 1185., (©2024 American Association for Cancer Research.)

The ability to monitor anti-tumor CD8+ T cell responses in the blood has tremendous therapeutic potential. Here, we used paired single-cell RNA and TCR sequencing to detect and characterize "tumor-matching" (TM) CD8+ T cells in the blood of mice with MC38 tumors or melanoma patients using the TCR as a molecular barcode. TM cells showed increased activation compared with nonmatching T cells in blood and were less exhausted than matching cells in tumors. Importantly, PD-1, which has been used to identify putative circulating anti-tumor CD8+ T cells, showed poor sensitivity for identifying TM cells. By leveraging the transcriptome, we identified candidate cell surface markers for TM cells in mice and patients and validated NKG2D, CD39, and CX3CR1 in mice. These data show that the TCR can be used to identify tumor-relevant cells for characterization, reveal unique transcriptional properties of TM cells, and develop marker panels for tracking and analysis of these cells., Competing Interests: Disclosures:   K. Tsai reported personal fees from Regeneron, grants from Array/Pfizer, grants from Replimune, and grants from Oncosec outside the submitted work. M. Rosenblum reported other from TRex Bio and other from Sitryx Bio outside the submitted work. A. Daud reported grants from Merck, grants from BMS, other from Trex, grants from Pfizer, grants from Incyte, grants from Xencor, grants from Roche, and grants from Novartis during the conduct of the study as well as grants from Oncosec outside the submitted work. A. Sharpe reported grants from NIH R01 CA229851, grants from NIH U54 CA224088, grants from NIH P01 56299, and grants from NIH P01 39671 during the conduct of the study; personal fees from Surface Oncology, personal fees from Sqz Biotech, personal fees from Selecta, personal fees from Monopteros, personal fees from Elstar, personal fees from Elpiscience, grants from Novartis, grants from Roche, grants from Merck, grants from Ipsen, grants from UCB, and grants from Quark Ventures outside the submitted work. In addition, A. Sharpe had a patent number 7,432,059 with royalties paid (Roche, Merck, Bristol-Myers-Squibb, EMD-Serono, Boehringer-Ingelheim, AstraZeneca, Leica, Mayo Clinic, Dako and Novartis), a patent number 7,722,868 with royalties paid (Roche, Merck, Bristol-Myers-Squibb, EMD-Serono, Boehringer-Ingelheim, AstraZeneca, Leica, Mayo Clinic, Dako and Novartis), a patent number 8,652,465 licensed (Roche), a patent number 9,457,080 licensed (Roche), a patent number 9,683,048 licensed (Novartis), a patent number 9,815,898 licensed (Novartis), a patent number 9,845,356 licensed (Novartis), a patent number 10,202,454 licensed (Novartis), a patent number 10,457,733 licensed (Novartis), a patent number 9,580,684 issued (none), a patent number 9,988,452 issued (none), and a patent number 10,370,446 issued (none); A. Sharpe is on the scientific advisory boards for the Massachusetts General Cancer Center, Program in Cellular and Molecular Medicine at Boston Children's Hospital, and the Human Oncology and Pathogenesis Program at Memorial Sloan Kettering Cancer Center and is a scientific editor for the Journal of Experimental Medicine. M. Singer reported personal fees from Guardant Health outside the submitted work. No other disclosures were reported., (© 2021 Pauken et al.)

Purpose: We have demonstrated previously that a mixture of three anti-Her-2 monoclonal antibodies (MAbs) that bind to different epitopes on the extracellular domain of Her-2 expressed on a human breast cancer cell line has more potent antitumor activity than the individual MAbs both in vitro and in xenografted severe combined immunodeficient mice. Because the activity of Herceptin is Fc dependent, we determined whether this would also be the case when a mixture of these three anti-Her-2 MAbs was used., Experimental Design: IgG and highly purified F(ab')(2) fragments of the anti-Her-2 MAbs and Herceptin were prepared and evaluated for their ability to induce cell death, inhibit vascular endothelial growth factor secretion, and mediate antibody-dependent cellular cytotoxicity and complement-mediated cytotoxicity in vitro. They were also compared for their abilities to induce regression of large BT474 tumors in severe combined immunodeficient mice., Results: All of the F(ab')(2) fragments were >95% pure and, as expected, did not mediate antibody-dependent cellular cytotoxicity or complement-dependent cytotoxicity in vitro. The in vitro antiproliferative and proapoptotic effects of the IgGs and F(ab')(2) fragments were similar. In contrast, the IgGs had significant antitumor activity in vivo, whereas their F(ab')(2) fragments were only marginally effective even at 5-fold higher doses to offset their shorter half-lives., Conclusions: These results confirm the importance of the Fc portion of Herceptin for optimal in vivo activity and demonstrate that even a mixture of three anti-Her-2 MAbs that are highly effective at inducing cell death in vitro requires Fc-mediated effector function for optimal in vivo activity.

Haemophilus influenzae can utilize different protein-bound forms of heme for growth in vitro. A previous study from this laboratory indicated that nontypeable Haemophilus influenzae (NTHI) strain N182 expressed three outer membrane proteins, designated HgbA, HgbB, and HgbC, that bound hemoglobin or hemoglobin-haptoglobin and were encoded by open reading frames (ORFs) that contained a CCAA nucleotide repeat. Testing of mutants expressing the HgbA, HgbB, and HgbC proteins individually revealed that expression of any one of these proteins was sufficient to allow wild-type growth with hemoglobin. In contrast, mutants that expressed only HgbA or HgbC grew significantly better with hemoglobin-haptoglobin than did a mutant expressing only HgbB. Construction of an isogenic hgbA hgbB hgbC mutant revealed that the absence of these three gene products did not affect the ability of NTHI N182 to utilize hemoglobin as a source of heme, although this mutant was severely impaired in its ability to utilize hemoglobin-haptoglobin. The introduction of a tonB mutation into this triple mutant eliminated its ability to utilize hemoglobin, indicating that the pathway for hemoglobin utilization in the absence of HgbA, HgbB, and HgbC involved a TonB-dependent process. Inactivation in this triple mutant of the hxuC gene, which encodes a predicted TonB-dependent outer membrane protein previously shown to be involved in the utilization of free heme, resulted in loss of the ability to utilize hemoglobin. The results of this study reinforce the redundant nature of the heme acquisition systems expressed by H. influenzae.

This study examined four commonly used sports knee evaluation instruments to establish normative data. A total of 91 volunteers who had not sought treatment for knee pain or any other knee complaint at any time in the past underwent a detailed medical history and examination of both of their knees. Three numerical systems (Feagin and Blake Knee Score, Lysholm Knee Score, and the Hospital for Special Surgery Knee Disability Assessment) and one nonnumerical system (the International Knee Documentation Committee Quick Knee Profile) were evaluated. A significant difference in the proportion of knees with excellent/normal ratings was seen among the four evaluation systems (P<.0001): Hospital for Special Surgery Knee Disability Assessment, 99.5%; Feagin and Blake Knee Score, 84.6%; Lysholm Knee Score, 84.1%; and the International Knee Documentation Committee Quick Knee Profile, 50.5%. Results showed that three objective component scores for the International Knee Documentation Quick Knee Profile were significantly lower than all other component scores (P<.05). These components included: overall ligament examination, Lachman, and total AP translation. These data may be useful as a baseline by which investigators studying patients following knee reconstructive procedures have a basis for comparison of their results.

This study investigated the incidence and clinical significance of postoperative fever in 118 consecutive patients undergoing 141 total knee arthroplasties (TKAs). A postoperative fever was recorded in 63 (66%) of 95 unilateral and 17 (74%) of 23 bilateral TKA patients. Nine of the unilateral and five bilateral TKA patients developed positive clinical or laboratory findings to explain the pyrexia. Unilateral TKA patients who experienced postoperative fever were statistically more likely to have a complication in the immediate postoperative period. None of the surgical variables examined had any predictive value on the incidence of postoperative fever. Aggressive pulmonary toilet, repeated physical examinations, and urine analysis are recommended when evaluating TKA patients with postoperative fever. Fever following TKA was common and was not necessarily a contraindication to discharge.