Combined Therapeutic Regimen Research Articles

Sequential doxycycline and colchicine combination therapy in Covid-19: The salutary effects

Coronavirus virus disease 2019 (COVID-19) is a viral infectious disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), actually considered as a global pandemic. The entry-point for SARS-CoV-2 is angiotensin converting enzyme 2 (ACE2) and dipeptidyl peptidase 4 (DPP4), which are highly expressed in the lung. Among other complications, COVID-19leads to fatal pneumonia, acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) due to development of cytokine storm (CS). The pathogenesis of SARS-CoV-2 infection depends on the viral load and human innate/adaptive immune response that are required for viral elimination in the first phase of COVID-19. However, an exaggerated immune response in the second phase of COVID-19 results in immune overreaction and CS-induced ALI and ARDS. Thus, in view of these considerations, we report here a series of five patients with COVID-19 pneumonia who developed ALI. In addition to the supportive therapy, the patients received doxycycline in the first week and doxycycline plus colchicine in the second week. Following sequential therapy with doxycycline and/or colchicine in patients with COVID-19 pneumonia, the patients had reduction of disease severity and symptoms with better clinical and radiological outcomes. However, it is tough to confirm the link between this therapeutic combination and recovery from COVID-19 pneumonia, as it is a small case-series report. Nevertheless, this study gives a rational for large-scale prospective studies to evaluate the dual sequential effect of doxycycline and colchicine on the COVID-19 severity. This case-series illustrated that use of colchicine: doxycycline combination is linked with marked improvements in the clinical, laboratory and radiological outcomes in patients with COVID-19 pneumonia. However, we cannot sketch any definitive conclusion from our observation, despite we hypothesize that this combination therapeutic regimen may attenuate and treat COVID-19. Further, namely prospective, randomized, and controlled clinical studies are recommended in this regard.

Impact of a diagnostic therapeutic educational pathway program for asthma management in preschool children

BackgroundPreschool children with clinically-diagnosed asthma have a higher rate of emergency department visits and consume more resources for management than older children. However, no clinical trials have yet been performed measuring the impact of a combined diagnostic, therapeutic and educational pathway regimen for evaluation of wheezing control in children aged less than 6 years.The purpose of the present study was to assess the impact of a pediatric program developed in Italy, the Diagnostic Therapeutic Educational Pathway (DTEP), for asthma management in children less than 6 years old attending an asthma referral center.MethodsThis is a retrospective population-based cohort study performed in children with asthma aged 0–5 years, attending at “Io e l’Asma center”, Brescia, Italy between September 2007 and December 2014. The incidence rates (IRs) of hospitalization, emergency room visits, use of outpatient services and drug usage for dyspnea, wheezing, or respiratory symptoms were evaluated for time periods prior to and after DTEP intervention.ResultsA total of 741 patients, aged 0–5 years completed the DTEP, including 391 and 350 children aged 0–2 and 3–5 years, respectively. The percentage of children aged 0–2 and 3–5 years showing improved control of wheezing symptoms during the 1st to 3rd visit interval as a result of the DTEP intervention increased from 39.5 to 60.9% and from 25.5 to 75.5%, respectively. During these periods, the IRs showed a significant decrease for all outcomes, from-8.6% to − 80.4%. Although specific IRs for drug prescriptions declined, particularly for LABA plus corticosteroids, antibiotics, and systemic corticosteroids, they increased for SABA, inhaled corticosteroid and leukotriene receptor antagonist usage.ConclusionsThe results suggest that a real-world assessment of the integrated DTEP program for preschool children provides evidence for improved wheezing control and reduction of adverse therapeutic related outcomes.

GM-CSF: Anti-Cancer Immune Response and Therapeutic Application

A primary focus of cancer therapeutics today is precision, or target directed, therapy. Combination treatment with precision therapy can involve both immune and signal pathway targets. One approach involving the chemokine GMCSF involves enhancement of the immune system. Herein is a review of the literature and the current therapeutic role of GM-CSF, including the proposed immune mechanisms and potential applications of GM-CSF to enhance anticancer immunotherapy. GM-CSF’s potent effects on dendritic cell activation and subsequent stimulation of T-lymphocyte activity make it an attractive potential addition to combination therapeutic regimens, including radiation therapy, oncolytic viral therapy, immune checkpoint inhibition, and autologous tumor vaccines, and warrants further clinical exploration, with an emphasis on identifying concomitant molecular pathways that mediate resistance and sensitivity to the GM-CSF effect.

Accelerated Healing of Chronic Wounds under a Combinatorial Therapeutic Regimen Based on Cold Atmospheric Plasma Jet Using Contact and Noncontact Styles

Accelerated Healing of Chronic Wounds under a Combinatorial Therapeutic Regimen Based on Cold Atmospheric Plasma Jet Using Contact and Noncontact Styles

A therapeutic DNA vaccine and gemcitabine act synergistically to eradicate HPV-associated tumors in a preclinical model

ABSTRACT Although active immunotherapies are effective strategies to induce activation of CD8+ T cells, advanced stage tumors require further improvements for efficient control. Concerning the burden of cancer-related to Human papillomavirus (HPV), particularly the high incidence and mortality of cervical cancer, our group developed an approach based on a DNA vaccine targeting the HPV-16 E7 oncoprotein (pgDE7h). This immunotherapy is capable of inducing an antitumour CD8+ T cell response but show only partial control of tumors in more advanced growth stages. Here, we combined a chemotherapeutic agent (gemcitabine- Gem) with pgDE7h to overcome immunosuppression and improve antitumour responses in a preclinical mouse tumor model. Our results demonstrated that administration of Gem had synergistic antitumor effects when combined with pgDE7h leading to eradication of both early-stages and established tumors. Overall, the antiproliferative effects of Gem observed in vitro and in vivo provided an optimal window for immunotherapy. In addition, the enhanced antitumour responses induced by the combined therapeutic regimen included enhanced frequencies of antigen-presenting cells (APCs), E7-specific IFN-γ-producing CD8+ T cells, and cytotoxic CD8+ T cells and, concomitantly, less pronounced accumulation of immunosuppressive myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs). These findings demonstrated that the combination of Gem and an active immunotherapy strategy show increased effectiveness, leading to a reduced need for multiple drug doses and, therefore, decreased deleterious side effects avoiding resistance and tumor relapses. Altogether, our results provide evidence for a new and feasible chemoimmunotherapeutic strategy that supports future clinical translation.

Clinical evaluation of HIFU combined with GnRH-a and LNG-IUS for adenomyosis patients who failed to respond to drug therapies: two-year follow-up results

Objective To investigate the clinical effect of high intensity focused ultrasound (HIFU) ablation combined with gonadotropin-releasing hormone agonist (GnRH-a) and levonorgestrel-releasing intrauterine system (LNG-IUS) in the treatment of adenomyosis patients who failed to respond to drug therapies. Study design A total of 47 patients with adenomyosis who had failed to respond to drug therapies and had no fertility desires were treated with HIFU combined with GnRH-a and LNG-IUS. The score of dysmenorrhea and menstrual volume were measured at pre and 6-, 12-, 18-, 24-month post-HIFU. Results All patients completed HIFU ablation without major postoperative complications. Compared with the symptom scores before the HIFU treatment, the score of dysmenorrhea and menstrual volume decreased significantly at 6, 12, 18 and 24 months after HIFU treatment (p < 0.05), but no significant difference was observed between 6, 12, 18 and 24 months after HIFU (p > 0.05). The clinical success rate was 100%, 100%, 95.7% and 93.6% respectively at 6, 12, 18 and 24 months after the combined treatment. Conclusion The combined therapeutic regimen of HIFU, GnRH-a and LNS-IUS is safe and effective, which can be an alternative treatment option for patients with adenomyosis who failed to respond to drug therapies to avoid adenomyomectomy or hysterectomy.

Synergistic cytotoxicity of erianin, a bisbenzyl in the dietetic Chinese herb Dendrobium against breast cancer cells

Erianin (ER), a dietary compound extracted from Dendrobium, a traditional Chinese medicinal edible herb, is well recognized for its potential anti-cancer activity. Nevertheless, its limitations, regarding its complex isolation procedure, low yield and low water solubility, limit large scale application. Combinatorial therapeutic regimen that combines several drugs to target different pathways in a characteristically synergistic manner at lower doses of drugs proved effective in several diseases treatment. Besides, new knowledge aimed at improving drug delivery into the intracellular environment is essential. In this study, ER was assessed for its cytotoxic effect in combination with doxorubicin hydrochloride (DOX·HCl) against breast cancer cells. Drug synergy was calculated by using combination index (CI) index and we discovered that they had positive effects. To ensure uniform delivery of both drugs to cells for a desired synergistic action, a dual drug loaded liposomes was developed using thin-film dispersion, and coated by a layer of folate-chitosan. Cytotoxicity and cell proliferation based assays revealed the increase of cell inhibition rate by more than 30% compared with free drugs. Fluorescence imaging revealed that liposomes can aid faster drugs accumulate in cancer cells. The study presented a novel strategy for the treatment of breast cancer.

Evolution of Peptide-Based Prostate-Specific Membrane Antigen (PSMA) Inhibitors: An Approach to Novel Prostate Cancer Therapeutics.

Prostate cancer is one of the most common cancers worldwide, with approximately 1.1 million cases diagnosed annually. The rapid development of molecular imaging has facilitated greater structural understanding, which can help formulate novel combinations of therapeutic regimens and more accurate diagnosis, avoiding unnecessary prostate biopsies. This accumulated knowledge also provides a greater understanding of the aggressive stages of the disease and tumor recurrence. Recently, much progress has been made on developing peptidomimetic-based inhibitors as promising candidates to effectively bind to the prostate- specific membrane antigen (PSMA), which is expressed by prostate cancer cells. In this review, recent advances covering small-molecule and peptide-based PSMA inhibitors will be extensively reviewed, providing a base for the rational design of future PSMA inhibitors. Herein, the literature on selected PSMA inhibitors that have been developed from 1996 to 2020 were reviewed, emphasizing recent synthetic advances and chemical strategies whilst highlighting therapeutic potential and drawbacks of each inhibitor. Synthesized inhibitors presented in this review demonstrate the clinical application of certain PSMA inhibitors, exhibited in vitro and in vivo. This review highlights the clinical potential of PSMA inhibitors, analyzing the advantages and setbacks of the chemical synthetic methodologies utilized, setting precedence for the discovery of novel PSMA inhibitors for future clinical applications.

Fatty Acid Oxidation Is an Adaptive Survival Pathway Induced in Prostate Tumors by HSP90 Inhibition

HSP90 is a molecular chaperone required for stabilization and activation of hundreds of client proteins, including many known oncoproteins. AUY922 (luminespib), a new-generation HSP90 inhibitor, exhibits potent preclinical efficacy against several cancer types including prostate cancer. However, clinical use of HSP90 inhibitors for prostate cancer has been limited by toxicity and treatment resistance. Here, we aimed to design an effective combinatorial therapeutic regimen that utilizes subtoxic doses of AUY922, by identifying potential survival pathways induced by AUY922 in clinical prostate tumors. We conducted a proteomic analysis of 30 patient-derived explants (PDE) cultured in the absence and presence of AUY922, using quantitative mass spectrometry. AUY922 significantly increased the abundance of proteins involved in oxidative phosphorylation and fatty acid metabolism in the PDEs. Consistent with these findings, AUY922-treated prostate cancer cell lines exhibited increased mitochondrial mass and activated fatty acid metabolism processes. We hypothesized that activation of fatty acid oxidation is a potential adaptive response to AUY922 treatment and that cotargeting this process will sensitize prostate cancer cells to HSP90 inhibition. Combination treatment of AUY922 with a clinical inhibitor of fatty acid oxidation, perhexiline, synergistically decreased viability of several prostate cancer cell lines, and had significant efficacy in PDEs. The novel drug combination treatment induced cell-cycle arrest and apoptosis, and attenuated the heat shock response, a known mediator of HSP90 treatment resistance. This combination warrants further preclinical and clinical investigation as a novel strategy to overcome resistance to HSP90 inhibition. IMPLICATIONS: Metabolic pathways induced in tumor cells by therapeutic agents may be critical, but targetable, mediators of treatment resistance.

Study protocol for an open-label, single-arm, phase Ib/II study of combination of toripalimab, nab-paclitaxel, and gemcitabine as the first-line treatment for patients with unresectable pancreatic ductal adenocarcinoma

BackgroundPancreatic ductal adenocarcinoma (PDAC) is a fatal disease with a dismal response to single-use of immune checkpoint inhibitors (ICIs). ICIs combined with systemic therapy has shown efficacy and safety in various solid tumors. Nab-paclitaxel and gemcitabine (AG), as the standard first-line treatment for advanced PDAC, has been widely used in recent years. The combination of ICIs and AG chemotherapy appears to be a promising option in the treatment of PDAC.MethodsThis is an open-label, single-arm, and single-center phase Ib/II trial. The enrolled subjects are the unresectable (locally advanced or metastatic) PDAC patients without previous systemic treatments. All subjects receive an intravenous injection of gemcitabine 1000 mg/m2 and nab-paclitaxel 125 mg/m2 on day 1 and day 8, along with toripalimab 240 mg at day 1 every 3 weeks. The subjects may discontinue the treatment because of progression disease (PD), intolerable toxicities, requirements of patients or researchers. For local advanced patients who are evaluated as partial response (PR), surgeons need to assess the surgical possibility. The primary objective of this trial is to evaluate the safety and overall survival (OS) of this combination therapy; and the secondary objective is related to the assessment of objective response rate (ORR), disease control rate (DCR), progression-free survival (PFS), and the rate of resection or R0 resection after receiving toripalimab plus AG treatment. Besides, we expect to identify the predictive biomarkers (such as MMR protein and PD-L1 expression, the number of TILs, the small RNA of EBV and so on) and explore the correlation between these biomarkers and tumor response to this combined regimen.DiscussionThis trial is the first attempt to evaluate the efficacy and safety of the combination of toripalimab plus AG chemotherapy as a first-line treatment for unresectable PDAC patients. The results of this phase Ib/II study will provide preliminary evidence for further assessment of this combined therapeutic regimen for unresectable PDAC patients.Trial registrationTrial registration: ChiCTR (ChiCTR2000032293). Registered 25 April 2020 - Retrospectively registered.

In Situ Forming Nutritional and Temperature Sensitive Scaffold Improves the Esthetic Outcomes of Meshed Split-Thickness Skin Grafts in a Porcine Model.

Objective: Full-thickness burn wounds require immediate coverage, and the primary clinical approaches comprise of skin allografts and autografts. The use of allografts is often temporary due to the antigenicity of allografts. In contrast, the availability of skin autografts may be limited in large burn injuries. In such cases, skin autografts can be expanded through the use of a skin mesher, creating meshed split-thickness skin grafts (MSTSGs). MSTSGs have revolutionized the treatment of large full-thickness burn injuries since the 1960s. However, contractures and poor esthetic outcomes remain a problem. We previously formulated and prepared an in situ forming skin substitute, called MeshFill (MF), which can conform to complex shapes and contours of wounds. The objective of this study was to assess the esthetic and wound healing outcomes in full-thickness wounds treated with a combination of MF and MSTSG in a porcine model. Approach: Either MSTSGs or MSTSG+MF was applied to full-thickness excisional wounds in Yorkshire pigs. Wound healing outcomes were assessed using histology, immunohistochemistry, and wound surface area analysis from day 10 to 60. Clinical evaluation of wounds were utilized to assess esthetic outcomes. Results: The results demonstrated that the combination of MSTSGs and MF improved wound healing and esthetic outcomes. Innovation: Effects of MSTSGs and reconstitutable liquid MF in a full-thickness porcine model were investigated for the first time. Conclusion: MF provides promise as a combination therapeutic regimen to improve wound healing and esthetic outcomes.

Combination of Chinese and western medicine to prevent and reverse resistance of cancer cells to anticancer drugs

Cancer is a common and complex disease with various genetic and environmental risks. Chemotherapy and targeted therapy are one of the main therapeutic approaches for the treatment of advanced cancer patients. However, prolonged patient treatment often provokes drug resistance phenomena that render the therapeutic treatment highly ineffective. Therefore, the development of novel and effective treatment strategies to overcome drug resistance is urgent for cancer therapy. A variety of factors contribute to drug resistance, including tumor cell heterogeneity, drug efflux, drug inactivation, changes in tumor microenvironment, DNA damage repair disorders, epigenetic alterations in cancer cells and so on. However, how to circumvent drug resistance to improve anticancer efficacy remains to be determined. Currently, the most important approach for overcoming drug resistance is drug combination therapeutic regimen, however, it is easy to induce new drug resistance or even multi-drug resistance (MDR) which is defined as the resistance of cancer cells to multiple anticancer drugs with different structures and mechanisms of action. Whereas, the “multi-components and multi-targets” manifestations of Traditional Chinese Medicine (TCM) make them an attractive direction for developing new adjuvant therapeutics to reverse drug resistance. In recent years, several compounds derived from TCM have been reported to augment sensitivity or overcome resistance of anticancer therapeutic drugs, including elemene, bufalin, curcumin, magnolol, honokiol, resveratrol, quercetin, and so on. Thus, the combination of TCM and conventional anticancer drugs will hopefully provide a novel strategy for enhancing the effects of anticancer strategies and overcoming their resistance in the treatment of cancer patients. In the current paper, we review the molecular mechanisms of drug resistance and the therapeutic strategies. We also discuss the current basic research and clinical studies in overcoming drug resistance from Traditional Chinese Medicine. Finally, we focus the integrated traditional Chinese and Western medicine as a promising new therapeutic approach to enhance the effects of anticancer strategies or overcome their resistance in the treatment of cancer patients.

The Fight Against Obesity Escalates: New Drugs on the Horizon and Metabolic Implications.

There is currently a steep rise in the global prevalence of obesity. Pharmaceutical therapy is a valuable component of conservative obesity therapy. Herein, medications currently in the phase of preclinical or clinical testing are reviewed, along with an overview of the mechanisms that regulate energy intake and expenditure. In addition, the current and potential future directions of obesity drug therapy are discussed. Although the current arsenal of obesity pharmacotherapy is limited, a considerable number of agents that exert their actions through a variety of pharmacodynamic targets and mechanisms are in the pipeline. This expansion shapes a potential near future of obesity conservative management, characterized by tailored combined therapeutic regimens, targeting not only weight loss but also improved overall health outcomes. The progress regarding the elucidation of the mechanisms which regulate the bodily energy equilibrium has led to medications which mimic hormonal adaptations that follow bariatric surgery, in the quest for a "Medical bypass." These, combined with agents which could increase energy expenditure, point to a brilliant future in the conservative treatment of obesity.

CYP3A7 drug‐hormone Interactions in the Neonate: Adrenal Insufficiency and Low Birthweight in the HIV infected Neonate caused by Inhibition of CYP3A7 DHEA‐S Oxidation by Ritonavir

In the neonate and developing infant, CYP3A7 is the predominant CYP3A isoform and is responsible for the majority of metabolism of CYP3A substrates. However, its primary function is the 16α‐hydroxylation of the endogenous hormone DHEA‐S, which is important for maintenance of hormone homeostasis in the adrenal gland and production of estriol during the third trimester of pregnancy. Low maternal estriol levels have been reported to lead to low birth weight and premature birth. It has been previously reported that cohorts of HIV infected mothers and their newborns on a combined therapeutic regimen of Lopinavir and Ritonavir have increased adrenal insufficiency and low birth weight, as well as substantially elevated levels of DHEA‐S, leading to adverse outcomes in this fragile patient group. Since Ritonavir is a known potent inhibitor of CYP3A4, we hypothesized that it might also be an inhibitor of CYP3A7‐mediated DHEA‐S oxidation and that this inhibition might be responsible for the adrenal insufficiency and low birth weight associated with Lopinavir‐Ritonavir treatment. In order to test this hypothesis, we employed both recombinant CYP enzyme and neonatal human liver microsomes (nHLMs). We observed significant inhibition of CYP3A7 DHEA‐S hydroxylation activity in the presence of Ritonavir with an IC50 of 0.05 μM. Furthermore, Ritonavir displayed time‐dependent inhibition of DHEA‐S oxidation, with a Ki of 0.336 μM and a Kinact of 0.1077 min−1, suggesting that it may be a mechanism based inactivator of CYP3A7. Lopinavir was also capable of inhibiting CYP3A7, albeit to a much lesser extent than Ritonavir with an IC50 of 5.88 μM for DHEA‐S 16α‐hydroxylation, and without the associated time‐dependency. Interestingly, we observed that Lopinavir alone had a lower intrinsic clearance (Clint) rate in nHLMs compared to adult HLMs, suggesting that a pharmacokinetic enhancer adjuvant may not be necessary in this patient population in order to slow drug clearance and maintain efficacy. At a minimum, our results suggest that closer monitoring of the effective dose of the pharmacokinetic enhancer in neonatal HIV therapy may be required in order to prevent significant inhibition of CYP3A7 DHEA‐S oxidation and the associated adverse effects.Support or Funding InformationThis work was supported by the National Institutes of Health, Institute of General Medical Sciences [Grant R01 GM128508‐03] to JNL.

Cellular rewiring in lethal prostate cancer: the architect of drug resistance.

Over the past 5 years, the advent of combination therapeutic strategies has substantially reshaped the clinical management of patients with advanced prostate cancer. However, most of these combination regimens were developed empirically and, despite offering survival benefits, are not enough to halt disease progression. Thus, the development of effective therapeutic strategies that target the mechanisms involved in the acquisition of drug resistance and improve clinical trial design are an unmet clinical need. In this context, we hypothesize that the tumour engineers a dynamic response through the process of cellular rewiring, in which it adapts to the therapy used and develops mechanisms of drug resistance via downstream signalling of key regulatory cascades such as the androgen receptor, PI3K-AKT or GATA2-dependent pathways, as well as initiation of biological processes to revert tumour cells to undifferentiated aggressive states via phenotype switching towards a neuroendocrine phenotype or acquisition of stem-like properties. These dynamic responses are specific for each patient and could be responsible for treatment failure despite multi-target approaches. Understanding the common stages of these cellular rewiring mechanisms to gain a new perspective on the molecular underpinnings of drug resistance might help formulate novel combination therapeutic regimens.

Abstract A37: Blockading PD1 on tumor-primed CD4 T cells instigates antitumor immunity

Abstract Despite the remarkable treatment benefits associated with interventional checkpoint PD1 blockade in cancer, most patients ultimately progress on therapy. Revealing mechanisms underlying tumors responsive to PD1 blockade is critical to identifying biomarkers for predicting responses to PD1 therapy and developing novel PD1 combined therapeutic regimens for their broad use in clinic. In a clinically reflective mouse model of spontaneous metastatic breast carcinoma, effective and durable PD1 therapy is dependent on systemic immunity including Type 1 conventional dendritic cells (cDC1: CD8α+DC/CD103+DC), B cells, CD4 T cells, and CD8 T cells. Increased circulating and/or tumor-infiltrating IL12+CD103+DC and IL2+CD4/CD8 T cells are associated with tumors responsive to PD1 therapy. PD1 blockade results in tumor-primed CD44+IL2+CD4 T cells in the periphery regardless of cDC1. Adoptive transfer of PD1 blockade-driven CD44+CD4 T cells ignites protective immunity against new tumors. Neutralization of IL2 post the adoptive transfer abrogates the antitumor immunity. These data suggest the prominent role of PD1 blockade on tumor-primed CD4 T cells in newly induced T cells via cDC1 in response to PD1 therapy. Citation Format: Xingxing Hao, Louis D. Falo III, Guo Chen, Cara D. Carey, Louis D. Falo Jr., Zhaoyang You. Blockading PD1 on tumor-primed CD4 T cells instigates antitumor immunity [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr A37.

Abstract P5-04-03: Conversion of non-T cell-inflamed triple-negative breast cancer by in situ immunomodulation of induced intratumoral cross-presenting dendritic cells

Abstract Checkpoint inhibitor (CPI)-based immunotherapy such as PD-1/PD-L1 blockade has shown an unprecedented success for the treatment of a variety of malignancies; however, meaningful clinical activity is seen only in a subset of breast cancer patients. This is due, at least in part, to poor T-cell infiltration into the tumor prior to CPI therapy, which is negatively correlated with the treatment response (non-T cell-inflamed tumor). Effective priming and trafficking of tumor-specific T cells to the tumor microenvironment (TME) can theoretically overcome current limitations of immunotherapy for non-T cell-inflamed tumors such as breast cancer. Accumulating evidence indicates that tumor-residing Batf3-dependent CD103+ dendritic cells (DCs) play critical roles in not only priming of tumor-specific CD8+ T cells, but also trafficking of CD8+ T cells to the TME, thus enhancing responses to PD-1/PD-L1 blockade. In the present study, we hypothesized that mobilization and activation of cross-presenting CD103+ DCs in the TME would increase CD8+ T cell infiltrate to the tumor, and facilitate effective regression of breast cancer. To this end, we utilized two triple-negative syngeneic mammary cancer models, 4T1 on a H-2d background and AT3 derived from the PyMT-MMTV model on a H-2b background, and evaluated antitumor efficacy of a combinatorial therapeutic regimen comprised of intratumoral administration of Fms-like tyrosine kinase 3 receptor ligand (Flt3L) to mobilize CD103+ DCs to the TME, local radiotherapy (RT) to induce immunogenic cell death and maturation of DCs, and peritumoral administration of a toll-like receptor 3 (TLR3) agonist to activate DCs. We found intratumoral administration of Flt3L increased the number of CD103+ DCs in the tumor, blood and lymph nodes, and local RT induced upregulation of TLR3, CD40 and CD86 on the tumor-residing CD103+ DCs. Following intratumoral administration of Flt3L, local RT, and in situ delivery of a TLR3 agonist significantly increased CD4+ and CD8+ tumor-infiltrating lymphocytes (TILs), mediated effective regression of large established tumors, and significantly improved survival in multiple mouse tumor models. Phenotypic analysis of circulating T cells revealed that the combinatorial regimen increased the frequency of effector memory and central memory T cell subsets with upregulation of PD-1 and C-X3-C motif chemokine receptor 1 (CX3CR1), suggesting effective activation and differentiation of T cells. Interestingly, we found the frequency of macrophages significantly decreased in the tumor after combinatorial treatment, indicating remodeling of both the myeloid and lymphoid compartments in the TME. Furthermore, increased PD-1 expression on CD4+ and CD8+ TILs, and PD-L1 expression in the TME suggest that antitumor efficacy of the combinatorial regimen can be additionally enhanced by PD-1/PD-L1 blockade. Taken together, combinatorial in situ immunomodulation with intratumoral Flt3L administration, RT and, peritumoral administration of a TLR3 agonist can convert non-T cell-inflamed tumors to T-cell inflamed tumors, remodel an immunosuppressive TME, and potentially overcome primary resistance to PD-1/PD-L1 blockade for triple-negative breast cancer. Acknowledgement: This work was supported by Uehara Memorial Foundation (T.O.), NIH/NCI K08CA197966 (F.I.) and P30CA016056 involving the use of Roswell Park Comprehensive Cancer Center’s Flow Cytometry Core. Citation Format: Takaaki Oba, Tibor Keler, Henry C Marsh, Scott Abrams, Fumito Ito. Conversion of non-T cell-inflamed triple-negative breast cancer by in situ immunomodulation of induced intratumoral cross-presenting dendritic cells [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P5-04-03.

Pulmonary-Affinity Paclitaxel Polymer Micelles in Response to Biological Functions of Ambroxol Enhance Therapeutic Effect on Lung Cancer.

PurposeCancer chemotherapy effect has been largely limited by cell autophagy and little drug accumulation at the action sites. Herein, we designed an intelligent strategy involving paclitaxel (PTX) polymer micelles in response to biological functions of ambroxol (Ax). The amphiphilic polymers polyethyleneglycol-polylactic acid (PEG-PLA) and Pluronic P105 were selected as nanocarriers to encapsulate PTX to form into lung affinity PEG-PLA/P105/PTX micelles. Ax which can up-regulate the secretion of pulmonary surfactant (PS) and inhibit autophagy was hired to change the microenvironment of the lung, thereby promoting the lung accumulation and increasing cell-killing sensitivity of the micelles.MethodsThe physical and chemical properties of the micelles were characterized including size, morphology, critical micellar concentration (CMC) and in vitro drug release behavior. The therapeutic effects of the combination regimen were characterized both in vitro and in vivo including study on Ax in promoting the secretion of pulmonary surfactant, in vitro cytotoxicity, cellular uptake, Western blotting, in vivo biodistribution, in vivo pharmacokinetics and in vivo antitumor efficacy.ResultsThe PEG-PLA/P105/PTX micelles showed a particle size of 16.7 ± 0.5 nm, a nearly round shape, small CMC and sustained drug release property. Moreover, the in vitro results indicated that Ax could increase PS and LC3 protein secretion and enhance the cytotoxicity of PEG-PLA/P105/PTX micelles toward A549 cells. The in vivo results indicated that the combination therapeutic regimen could promote the micelles to distribute in lung and enhance the therapeutic effect on lung cancer.ConclusionThis multifunctional approach of modulating the tumor microenvironment to enhance drug transportation and cell-killing sensitivity in the action sites might offer a new avenue for effective lung cancer treatment.

The in vivo modulatory effects of Cornus mas extract on photodynamic therapy in experimental tumors.

Photodynamic therapy (PDT) is a treatment of cancer due to its ability to induce cell death, oxidative stress and acute inflammatory reaction in targeted sites. To optimize the effect of PDT the addition of some compounds with supplementary cytotoxic effect on tumor cells was tried. The study was performed on 35 Wistar male albino rats with Walker 256 carcinosarcoma. The animals were randomly assigned in seven groups (n = 5) and treated as follows: group 1 - control; group 2 - Cornus mas (CM) extract 15 mg/kg b.w., administered for 7 days; group 3 - CM extract administered for 7 days followed by irradiation (CM + IR); group 4 - one dose of tetra-p-sulfonato-phenyl-porphyrin (TSPP) 10 mg/kg b.w.; group 5 - TSPP + IR; group 6 - CM extract administered daily for 7 days before TSPP and IR (CM + TSPP + IR); group 7 - TSPP + IR followed by CM administered for 7 days (TSPP + IR + CM). The results showed that MDA and GSSG levels increased after PDT in parallel with the increasing of COX-2 expression and DNA damage. Apoptotic and necrotic index enhanced in TSPP + IR, effect improved by CM association before PDT. CM + TSPP + IR regimen also induced more intense inflammatory reactions, increased COX-2 expression, determined DNA damage, apoptosis and necrosis, compared to the TSPP + IR + CM group. Both combined therapeutic regimens reduced MDA levels in tumor tissue, especially CM + TSPP + IR and increased the antioxidant defense and iNOS expression. Our results demonstrated that CM associated before PDT had beneficial effects in PDT and may represent a promising option in PDT strategies.

Checkpoints Under Traffic Control: From and to Organelles.

Immune checkpoints are variegated stimulatory and inhibitory signals that are fundamental in immune homeostasis. The regulative molecules for immune checkpoints include programmed cell death protein 1 (PD1), programmed death-ligand 1 or 2 (PD-L1 or PD-L2), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), and so on. While the immune checkpoint molecules have gained soaring attention in recent years, the trafficking of them has been rarely studied. Since all of the discovered immune checkpoint molecules are transmembrane domain (TMD) proteins, they share similar pathophysiological characteristics which make studies about their trafficking and associated disorders resembled. PD-L1 is one of the most classic immune checkpoint molecules, and anti-PD1 monoantibodies have shown promising immunotherapeutic effects. PD-L1 trafficking has been particularly studied, the key regulators of which include metformin, chemokine-like factor-like MARVEL transmembrane domain-containing family member (CMTM), Huntingtin-interacting protein 1-related (HIP1R), exosomes, ALIX, polyI:C, and various post-translational modifications. Here, we focus on the checkpoints under traffic control, counting PD-L1, CTLA-4, lymphocyte-activation gene 3 (LAG-3), killer immunoglobulin-like receptors (KIRs), CD70, CD94, and attempt to shed light on the potentials of drug targets based on these findings and look forward to further studies in combinatorial therapeutic regimens in the meantime.