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Dear Editor,In the recent publication titled ‘Search for tumor-specific frequencies of amplitude-modulated 27 MHz electromagnetic fields in mice with hepatocarcinoma xenografted tumors’, Veyret et ...

Pituitary tumors develop within a macroenvironment and microenvironment that favor their overall survival. Given the pituitary gland's unique anatomic, histological, and physiological properties, the role of the TME in the pituitary tumors is central. This chapter summarizes the body of literature investigating the role of each component of the TME in relation to tumors of the pituitary and its intimate association with tumorigenesis, maintenance, and progression.

Cancer cells divide continuously without any control, rendering immortal cells. These changes cause the dynamic instability in their genomes. Therefore each cancer cell in a tumor mass has various changes for specific tumor types. Their differences should be diagnosed on the basis of mutated genes and/or proteins. After the detection of alterations in cancer cells based on the techniques and instruments of biomedicine, personalized therapy continues with preexisting drug implementations. Personalized therapy broadens the borders with circulating tumor cells causing the metastasis in distant regions, developing specific gene therapies, novel targeted drugs, and immunotherapeutics for eliminating cancerous cells precisely. Within this context, biomedicine helps humankind to merge the biological and physiological knowledge to clinical practice. In this chapter, we aimed to clarify attitudes in biomedicine from molecular changes from single tumor cells to in vivo studies to find the best therapy for a cancer patient.

This chapter reviews the use of orthotopic mouse models and novel tumor-specific antibodies conjugated to near-infrared dyes for high-resolution detection of colon cancer and metastases. Fluorescence imaging can detect early colon cancer with colonoscopy and provide improved detection of tumor margins, sentinel nodes, and identification of occult metastases during surgery. We also detail emerging clinical applications of tumor-specific fluorescence imaging in colorectal cancer.

International audience; Aim: The Pasche research group has reported that tumor-specific electromagnetic field frequencies have physiological and potential anti-tumor effects in cells, animals, and humans. Our aim was to investigate whether these fields have similar effects on physiological parameters in murine tumor models. Methods: Human HuH7 or HEPG2 cells were implanted in the right flank of 8-week-old female RAG gamma 2C immunodeficient mice. An oximeter was used to record systolic blood pressure (pulse) in free-roaming conscious mice. Mice pulses were recorded and analyzed using a in-house software that also controlled the low-frequency generator for modulating the 27.12 MHz carrier wave at selected frequencies. Results: We performed exposures using both systematic scans at low-frequencies and at the predetermined frequencies reported by the Pasche group as altering both pulse and tumor growth in humans. Those exposures produced no detectable change in physiological parameters of tumor-bearing mice. Conclusion: No tumor-related frequencies were found, neither using systematic scans of frequencies nor published specific frequencies. There might obviously be differences between animal and human models, but our approach did not confirm the physiological data of the human Pasche group data.

Uptake of 18F-NaF is not tumor specific, and nonmalignant entities can demonstrate radiotracer uptake, including arthritis, trauma, and bone processes such as fibrous dysplasia and Paget disease. The degree of radiotracer uptake cannot be used to differentiate benign from malignant lesions. Many benign bone lesions, including osteophytes and degenerative endplate changes, will show increased 18F-NaF uptake [1].

Carbon quantum dots (Cdots) have attracted more and more interests in bioimaging and tumor theranostics. However, their practical application has been limited due to the small particle size and non-tumor-specific fluorescence. Here, reduction-cleavable disulfide-linked Cdots-based nanoclusters were fabricated to conjugate doxorubicin (DOX) via an acid-labile hydrazone bond. Owing to the pH and reduction dual-stimuli responsiveness, the proposed Cdots-based nanotheranostics possessed unique tumor-specific fluorescent property and tumor-specific controlled drug release performance, indicating their promising potential for the in-situ real-time fluorescent monitoring of therapeutic response in future tumor therapy.

Cancer immunotherapy continues to make headway as a treatment for advanced stage tumors, revealing an urgent need to understand the fundamentals of anti-tumor immune responses. Noteworthy is a scarcity of data pertaining to the breadth and specificity of tumor-specific T cell responses in metastatic breast cancer. Autochthonous transgenic models of breast cancer display spontaneous metastasis in the FVB/NJ mouse strain, yet a lack of knowledge regarding tumor-bound MHC/peptide immune epitopes in this mouse model limits the characterization of tumor-specific T cell responses, and the mechanisms that regulate T cell responses in the metastatic setting. We recently generated the NetH2pan prediction tool for murine class I MHC ligands by building an FVB/NJ H-2q ligand database and combining it with public information from six other murine MHC alleles. Here, we deployed NetH2pan in combination with an advanced proteomics workflow to identify immunogenic T cell epitopes in the MMTV-PyMT transgenic model for metastatic breast cancer. Five unique MHC I/PyMT epitopes were identified. These tumor-specific epitopes were confirmed to be presented by the class I MHC of primary MMTV-PyMT tumors and their T cell immunogenicity was validated. Vaccination using a DNA construct encoding a truncated PyMT protein generated CD8 + T cell responses to these MHC class I/peptide complexes and prevented tumor development. In sum, we have established an MHC-ligand discovery pipeline in FVB/NJ mice, identified and tracked H-2Dq/PyMT neoantigen-specific T cells, and developed a vaccine that prevents tumor development in this metastatic model of breast cancer.

Background: ALPPS is a two-stage hepatectomy that induces more rapid liver growth compared to conventional strategies. This report aims to establish a risk-score to avoid adverse outcomes of ALPPS only for patients with colorectal liver metastases (CRLM) as primary indication for ALPPS. Methods: All patients with CRLM included in the ALPPS registry were included. Risk score analysis was performed for 90-day mortality after ALPPS, defined as death within 90 days after either stage. Two risk scores were generated i.e. one for application before stage-1, and one for application before stage-2. Logistic regression analysis was performed to establish the risk-score. Results: In total, 486 patients were included, of which 35 (7%) died 90 days after stage-1 or 2. In the stage-1 risk score, age ≥67 years (OR 3.7), FLR/BW ratio 50 μmol/L (OR 2.4), and stage-1 morbidity grade IIIA or higher (OR 6.3) were included. Conclusions: The CRLM risk-score to predict mortality after ALPPS demonstrates that older patients with small remnant livers in inexperienced centers, especially after experiencing morbidity after stage-1 have adverse outcomes. The risk score may be used to restrict ALPPS to low-risk patient populations.

This article reviews technical issues to improve surgical safety and avoid surgical errors in pediatric surgical oncology, particularly in the three most common extracranial solid tumors: neuroblastoma, hepatoblastoma and Wilms tumor. The use of adjuvant chemotherapy - when indicated - the use of tumor specific classifications, adequate surgical planning, that may include the use of 3D printable models, improved surgical instruments and technology, and following surgical guidelines, would result in avoiding error, increased safety, and therefore in improved surgical outcomes.

3047 Background: A considerable fraction of lung cancer patients raise diagnostic challenges requiring invasive procedures with a certain risk of complications. Therefore, new diagnostic tools are of major interest. Aberrant methylation of the HOXA9 gene occurs in almost all malignant lung tumors and HOXA9 methylated DNA (meth-ctDNA) is shed into the circulation. The present study aimed at a prospective investigation of the possible diagnostic value of HOXA9 meth-ctDNA in bronchial lavage (BL). Methods: Patients enrolled were referred from the general practitioner suspecting lung cancer. The diagnostic package according to national guidelines includes chest and abdominal CT scan, bronchoscopy, relevant blood tests, and histopathological or cytological verification. Twelve ml liquid was collected at bronchoscopy for analysis of meth-ctDNA based on ddPCR technology according to our published method. The analysis was performed blinded to the clinical data and compared to the final diagnosis. Results: Eighty-nine patients were consecutively included from the 1 November 2018 to 31 January 2019. Fifty-six patients (62.9%) were diagnosed with lung cancer and 33 (37.1%) with a variety of benign diseases. Meth-ctDNA was found in 42/56 of the patients with a malignant tumor, sensitivity = 75.0% (95%CI=61.6-85.6%), whereas 31/33 of the patients without cancer were negative, specificity = 93.9% (95%CI= 79.8-99.3%). Table summarizes the results. The false negative samples were mainly from patients with peripheral tumors. The two false positive patients included one patient with Cryptogenic Organizing Pneumonia and one with unspecific nodule. Conclusions: The presence of meth-ctDNA in BL has a high sensitivity and specificity. If validated, the analysis represents a valuable adjunct in the diagnosis of lung cancer. Potentially, it could save the patients from numerous examinations with potential harmful risks and ensure a fast diagnosis. The relation between meth-ctDNA and final lung cancer diagnosis (N= 89). [Table: see text]

Breast cancer is the second leading cause of cancer mortality in women after lung cancer. About 70% of breast cancers are positive for estrogen receptor alpha (ERα) at the time of diagnosis, and they respond well to anti-estrogen therapies; however, 40-50% of them become refractory over time. Thus, identifying novel therapeutic targets is critical to treat this ever-evolving disease. Recent studies suggest that the genes transcribed from chromosome X (X-linked genes) are implicated in several cancers. The expression of these genes is tightly regulated and restricted to immune-privileged organs; however, many of them escape regulation and are aberrantly expressed in tumors. Interestingly, our genomic analysis suggests that several of these genes are located at putative gene deserts in the human genome. To date, nothing is known about their expression and role in hormone-refractory breast cancer. In this regard, we used an integrated genomic approach to identify breast cancer-specific X-linked coding and noncoding genes. Furthermore, by combining gene expression analysis from MCF-7/TAMR-1 (tamoxifen resistant) breast cancer cells, we generated a comprehensive list of 100 estrogen-regulated, tamoxifen-resistant X-linked genes. Of 100 genes that met our stringent filtering criteria, 20 of them are long noncoding RNAs (lncRNAs), localize directly to chromatin and robustly regulated by estrogen. Each of these lncRNAs has a unique expression pattern in normal human tissues and breast cancer. For example, recently annotated primate-specific lncRNA1476 (a.k.a. UEBC - Upregulated in ER+ Breast Cancer), found in ovary, uterus, and fallopian tubes, and is localized to chromatin. The UEBC is regulated by estrogen and/or tamoxifen, and its expression predicts poor outcome in ERα+ breast cancer patients. UEBC has now been fully annotated (transcription start and stop site, 5’ cap, polyA tail, and exon/intron structure), and cloned. Gene expression combined with genome-wide analysis of ERα recruitment at enhancers indicate that UEBC plays a critical role in the estrogen-dependent transcription. Collectively, our results suggest that UEBC is an excellent target with prognostic and therapeutic potential in hormone-refractory breast cancer. Supported by a grant from the Cancer Prevention and Research Institute of Texas

Background Genetic testing to identify succinate dehydrogenase (SDH) mutations in patients with head and neck paraganglioma (HNP) has been in clinical practice for more than a decade. However, the recurrence and metachronous tumor occurrence risks in surgically treated mutation-positive patients are not well studied. Methods Clinical and procedural details of consecutive patients who underwent excision for HNP from January 1996 to October 2016 were retrospectively reviewed. End points included recurrence, metachronous tumor detection, and mortality. Germline DNA was tested to identify mutations in SDHx genes. Patients were divided into three groups on the basis of genetic testing: group I, positive; group II, negative; and group III, unknown or offered but not tested. Results HNP was diagnosed in 268 patients, 214 (147 female; mean age, 47 years) included in this study. Directed genetic testing was performed in 68; mutations were detected in SDH in 47 (69%), a majority SDHD. In group I, 47 patients had 64 procedures for 81 tumors (52 carotid body tumors [CBTs]); 17 (36%) were bilateral, 7 (15%) multiple, 3 (6%) functional, and 7 (15%) malignant. Residual tumor in 10 was significant in 2, managed by radiation therapy and reoperation. Local recurrence was detected in 12 patients (25%) at a median of 8 years; 11 metachronous mediastinal and retroperitoneal paragangliomas were detected in 8 (17%) at a median of 13 years. Systemic metastases occurred in five (10%). Six patients (13%) had more than one recurrence. In group II, 21 patients had 22 procedures for 23 tumors, 17 CBTs. Two (9%) were bilateral and two (9%) malignant. Excision was complete in all with no recurrence or systemic metastasis at last follow-up. For group III, 146 patients underwent 153 procedures for 156 tumors, 95 CBTs; 7 (5%) were bilateral, 2 (1%) multiple, 8 (5%) functional, and 1 (0.6%) malignant. Local recurrence was detected in nine (6%) at a median of 9 years and metachronous HNP in three (2%) at a median of 5 years. Systemic metastases occurred in two (1%). Mortality was 4% in group I and 3% in group III, none procedure or tumor related. Group I (mutation positive) had 10-year overall, recurrence-free, and metachronous tumor-free survival rates of 93%, 69.4%, and 73%, respectively, lower than the other groups (P Conclusions Bilateral, functional, malignant, recurrent, and metachronous tumors are more common in SDH mutation-positive patients with HNP. Overall survival in patients with HNP is high. Metachronous tumors or local recurrences occur late, and long-term follow-up is necessary.

Tumor-specific activatable therapy has gained more and more attention due to its improved therapeutic efficacy and minimized side effects. However, most of these nanomedicines only utilized one stimulus as the switch of the therapeutic process, which may have the issue of low selectivity. Herein, we develop a “dual lock-and-key” type activatable nanotherapeutic platform (ABTS@PAH-CNts) for tumor-specific photothermal therapy (PTT). Such nanoplatform can only be activated under both H2O2 and acidic pH, thus achieving higher selectivity towards tumor than single stimulus-activated nanomedicine. The nanoplatform is prepared by adsorbing 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) onto the surface of ceria nanotubes. We demonstrate that ceria nanotubes have analogous peroxidase activity with natural horseradish peroxidase (HRP). Under both H2O2 and acidic pH, ceria nanotubes can catalyze ABTS into its oxidized form, showing enhanced near-infrared (NIR) absorption with good photothermal effect. ABTS@PAH-CNts shows good in vitro biocompatibility without laser irradiation, while has good anticancer efficiency under 808 nm laser irradiation. In addition, in vivo study indicates that the photothermal temperature of ABTS@PAH-CNts can reach up to 55 °C in the tumor site, while only can reach 40 °C for normal tissue, demonstrating its good tumor specificity. Our work not only develops a nanozyme-based “dual lock-and-key”-controlled PTT system, but also provides a nanoplatform for activatable theranostics.

IL-12 is a cytokine produced by antigen-presenting cells that increases T cell proliferation, IFN gamma mediated Th1 effector functions and T and NK cell cytotoxicity. While these effects generate potent anti-tumor immunity in mouse models, the high toxicity of IL-12 in cancer patients has limited its clinical utility. Potency attenuation and intratumoral cytokine localization may improve IL-12 tolerability, but these approaches can reduce efficacy or be limited by intratumoral delivery or tumor antigen expression. To improve both tolerability and efficacy of IL-12, we engineered IL-12Fc fusions with anti-IL-12 antibodies to block IL-12 potency. Using linkers designed to be cleaved by highly active intratumoral proteases, blocking antibodies are released specifically in the tumor microenvironment, thereby increasing intratumoral IL-12 activity. Single chain IL-12 was fused to one C-termini of an Azymetric™ Fc heterodimer. To the other Fc C-termini, an anti-IL-12 scFv was fused via a protease-cleavable linker in order to block IL-12 activity. In addition to antibody blockade, modifications of IL-12Fc fusions were used to further limit IL-12Fc activity. In vitro potency was determined by CD8T cell IFN gamma release. Anti-IL-12 scFv cleavage in the presence of recombinant enzyme or human tumor material was assessed by reducing CE-SDS and LC/MS, respectively. All IL-12Fc fusions were produced with favorable biophysical characteristics. Modified IL-12Fc molecules had up to 500x reduced potency, while antibody blocked IL-12Fc molecules had up to 100,000x reduced potency compared to IL-12Fc control. In vitro cleavage of the anti-IL-12 scFv from IL-12Fc recovered potency to that of the corresponding non-masked IL-12Fc molecules. Reducing CE-SDS confirmed that anti-IL-12 scFvs were cleaved from IL-12Fc in the presence of recombinant enzyme. Cleavage of anti-IL-12 scFvs also occurred when variants were incubated in pancreatic tumor cell supernatant or human tumor tissue lysate. These results indicate that antibody blockade and re-activation by protease cleavage is a promising strategy to localize the activity of IL-12 to the tumor microenvironment while potentially limiting off-tumor toxicities. Further modifying antibody blocked IL-12 has the potential to better fine tune therapeutic index. We are pursuing tumor-specific IL-12 fusions for clinical application in tumors with high intratumoral protease activity. Citation Format: Jennifer Leah Bishop, Ryan Blackler, Gesa Volkers, Maya Poffenberger, Irene Yu, Joel Smith, Akram Khodabandehloo, Sifa Arrafi, Desmond Lau, Liz Stangle, Leisa Stenberg, Patricia Zwierzchowski, Iulia Dude, David Douda, Grant Wickman, Jeff Proctor, Gerry Rowse, Laurence Madera, Genevieve Desjardins, Nicole Afacan, Stuart Barnscher, David Mills, Thomas Spreter, Surjit Dixit. Increasing the therapeutic index of IL12 by engineering for tumor specific protease activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1788.

Hematopoietic progenitor kinase 1 (HPK1) is an intracellular protein kinase that negatively regulates T cell signaling and proliferation. Upon T cell receptor (TCR) activation, active HPK1 phosphorylates the adaptor protein SLP76 in the TCR complex, recruiting the negative regulator 14-3-3 and targeting components of the TCR signaling complex for degradation. HPK1 thus limits the TCR signaling important for mounting an effective immune response against tumor cells. We are employing structure-guided drug design to develop potent small-molecule inhibitors of HPK1. Our compounds potently inhibit HPK1 in biochemical assays, reduce levels of phosphorylated SLP76 and concomitantly increase IL-2 production by Jurkat T cells. Importantly, our HPK1 inhibitors enhance cytokine production by human and mouse primary T cells above that observed with TCR activation alone. Treatment of mice with orally available HPK1 inhibitors results in increased activation of antigen-specific CD8+ T cells in vivo and decreased tumor growth as single agent and in combination with clinically relevant checkpoint inhibitor antibodies. Our work confirms the importance of HPK1 for T cell function and supports HPK1 as a promising next generation immuno-oncology target. Citation Format: George Katibah, Yamini Ohol, Cyril Bucher, Lavanya Adusumilli, Deepika Kaveri, Omar Robles, Michael Sun, Cynthia Cho, Heather Milestone, Rachel Ames, Scott Jacobson, Dan Nebalasca, Justy Gomez- Guagua, Jerick Sanchez, Molly Grandcolas, Steve Wong, Martin Brovarney, Chandru Ramana, Thant Zaw, Lan Nguyen, Parcharee Tivitmahaisoon, Andrew Ng, Anqi Ma, Blanca Gomez, Michelle Ko, Paul Leger, Jeffrey Jackson, Grant Shibuya, Anton Shakhmin, Delia Bradford, Mengshu Xu, Mikhail Zibinsky, Daniel Poon, David Wustrow, Paul Kassner, Dirk Brockstedt. Development of small-molecule HPK1 inhibitors to unleash tumor-specific T cell responses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1646.

Many novel immune-oncology biologics are limited in clinical utility by narrow therapeutic windows. One strategy to overcome this limitation relies on the engineering of ‘masks' that block antibody paratopes outside of the tumor microenvironment (TME). The PROTECT (PROgrammed Tumor Engagement & Checkpoint/Costimulation Targeting) platform is designed to employ the orthogonal mechanistic features of a multispecific design to increase the therapeutic window by limiting exposure and activity in peripheral tissues while focusing activities to the tumor. In particular, we aim to bring TME-specific activity and enhanced immune modulation in a single transferable, conditionally active design. To achieve this, we engineered the N-termini of antibody heavy and light chains with the fusion of IgV domains of commonly targeted immunomodulatory pairs, such as PD-1 and PD-L1, to sterically preclude binding of the antibody paratopes to their target tumor antigens. We demonstrate that this approach can effectively mask the antibody binding and activity for targets by 10-1000 fold, with recovery of binding and anti-tumor activity upon release of the immunomodulatory mask once proteolytically cleaved by a tumor-specific protease such as urokinase plasminogen activator (uPA). In addition, we also demonstrate that we can selectively cleave and remove one half of the immunomodulatory pair (e.g. PD-L1) from the antibody thereby creating an antibody fused to only PD-1. The resulting bispecific antibody can now co-engage both the tumor antigen (TA) and its counterpart checkpoint inhibitor (PD-L1) to confer additional antitumor activities. As a proof of concept, we showed that for an engineered anti-CD3/HER2 bispecific T-cell engager (TCE), incorporation of the PD-L1/PD-1 PROTECT design creates a conditionally activated anti-CD3/PD-L1/HER2 tri-specific TCE that has (i) masked CD3 engagement with a reduction of the EC50 by 2 orders of magnitude, and (ii) enhanced T-cell dependent cytotoxicity over the parent anti-CD3/HER2 TCE by an order of magnitude once activated by uPA. Importantly, the enhanced activity of the tri-specific TCE is greater than the combination of the parental TCE with the anti-PD-L1 monoclonal antibody atezolizumab. We have also demonstrated that the PROTECT platform can be combined with anti-TA antibody to introduce a masking effect while increasing antibody-dependent cell cytotoxicity (ADCC) activity upon co-engagement of TA and PD-L1, again synergistic relative to the combination of anti-TA antibody and atezolizumab. Taken together, the PROTECT platform represents a novel approach to integrate immune modulation while limiting off-tumor activities for the development of conditional multispecific antibodies with potentially enhanced therapeutic window and activity. Citation Format: Surjit Dixit, Florian Heinkel, Anna Von Rossum, Harsh Pratap, Sifa Arrafi, Javairia Rahim, Purva Bhojane, Liz Stangle, Leisa Stenberg, Gesa Volkers, Eric Escobar-Cabrera, Thomas Spreter. PROTECT, a novel antibody platform for integrating tumor-specific immune modulation and enhancing the therapeutic window of targeted multispecific biologics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 924.

Tumor-specific mutated proteins can create non-self, mutation-containing ‘neoepitopes' that are immunogenic and are attractive targets for adoptive T-cell therapies. To avoid the complexity of patient-specific, private neoepitopes and their T-cell responses, there has been major interest in targeting common shared mutations in driver genes using off-the-shelf T-cell receptors engineered into autologous lymphocytes. The diversity of MHC alleles which could present these select neoantigens is an obstacle to this strategy. Most neoepitopes will not be successfully presented by most HLA alleles, and algorithms predicting epitope-HLA binding will either miss valid epitopes or identify too many candidate alleles to investigate thoroughly. One method to definitively demonstrate that an MHC allele can successfully present an epitope is to elute peptides bound to that MHC allele on the cell surface and analyze them by mass spectrometry (MS). It has the additional advantages of defining the precise epitope processed from an antigen and estimating its abundance. We undertook a comprehensive analysis by MS of several common shared mutations in driver oncogenes and their presentation by common Class I HLA alleles. To overcome the low sensitivity of MS (compared to T-cells), we created mono-allelic cell lines expressing one Class I HLA allele and one common mutated oncogene. Peptides bound to MHC on the surface of these cell lines were immunoprecipitated, eluted, and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). As expected, most HLA alleles could not present a detectable neoepitope from the candidate antigens. Our system was able to identify precise neoepitopes derived from four commonly mutated oncogenes (KRAS, EGFR, BRAF, and PIK3CA) presented by the HLA-A*03 HLA superfamily. The majority of mutated peptides were consistently found in biological replicates, demonstrating the reproducibility of our system. By evaluating intensities and fragmentation data for mutated peptides, we were able to improve the reliability of detection of neoepitopes at the cell surface. Although most of the identified epitopes were predicted to bind using the canonical NetMHCpan algorithm, many epitopes predicted to bind with high affinity were not observed using our pipeline. HLA-A*0201 has been intensely studied because of its high frequency in the U.S. population but we have not found it to consistently present mutated peptides from our candidate oncogenes. Our data show that we could utilize a MS approach to demonstrate which shared oncogene-derived neoepitopes are unequivocally processed and presented by common HLA alleles, which will allow greater focused efforts on developing TCRs against these promising HLA-neoantigen pairs. Citation Format: Catherine M. Ade, Yue A. Qi, Sudipto Das, Ken-ichi Hanada, Tapan Maity, Xu Zhang, Thorkell Andresson, Udayan Guha, James C. Yang. A mass spectrometry survey of frequent HLA alleles successfully presenting common tumor specific mutations for immune recognition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 273.

The emergence of biomimetic nanotechnology has seen an exponential rise over the past decade with applications in regenerative medicine, immunotherapy and drug delivery. In the context of nanomedicines activated by near infrared (NIR) photodynamic processes (photonanomedicines; PNMs), biomimetic nanotechnology is pushing the boundaries of activatable tumor targeted nanoscale drug delivery systems. This review discusses how, by harnessing a unique collective of biological processes critical to targeting of solid tumors, biomimetic PNMs (bPNMs) can impart tumor cell specific and tumor selective photodynamic therapy-based combination regimens. Through molecular immune evasion and self-recognition, bPNMs can confer both tumor selectivity (preferential bulk tumor accumulation) and tumor specificity (discrete molecular affinity for cancer cells), respectively. They do so in a manner that is akin, yet arguably superior, to synthetic molecular-targeted PNMs. A particular emphasis is made on how bPNMs can be engineered to circumvent tumor cell heterogeneity, which is considered the Achilles’ heel of molecular targeted therapeutics. Forward-looking propositions are also presented on how patient tumor heterogeneity can ultimately be recapitulated to fabricate patient-specific, heterogeneity-targeting bPNMs.

2559 Background: Conventional dendritic cells subtype 1 (cDC1) play a vital role in the priming and expansion of tumor specific CD8+ T cells and their recruitment to tumor microenvironment (TME). However, cDC1s are often underrepresented in the TME. Systemic administration of Fms-like tyrosine kinase 3 ligand (FLT3L), a hematopoietic growth factor that binds to FLT3 on myeloid and lymphoid progenitor cells, leads to expansion of cDC1s in the periphery which can then be recruited into the TME. We hypothesize that FLT3 pathway stimulation using GS-3583, a FLT3 agonist Fc fusion protein, has the potential to promote T cell mediated anti-tumor activity. Methods: This was a first-in-human placebo-controlled study of GS-3583 in healthy volunteers to evaluate the safety, PK, and PD of escalating single doses (ranging from 75μg to 2000μg) of GS-3583. The study was blinded to the subjects and the investigator. Each dose cohort enrolled 8-12 healthy subjects who received GS-3583 or placebo as single IV infusion at 3:1 ratio. Subjects were observed in the phase 1 unit for 15 days and then for 12 weeks as outpatients. As part of the PD evaluation, we investigated the changes in the number of cDC1s and cDC subtype 2 (cDC2) cells. Results: As of 8th Feb 2021, selected safety, PK and PD data from the first 3 cohorts were available. GS-3583 was well tolerated and all subjects had been discharged. To date, there have been no serious or grade 3 or higher adverse events. Preliminary PK analysis suggested dose-dependent increase in GS-3583 exposure (AUC and Cmax). Preliminary PD analysis shows that administration of GS-3583 resulted in dose-dependent increases in cDC1/cDC2 cells that peaked at day 5 or day 8 and returned to baseline within three weeks of drug administration. Conclusions: GS-3583 was safe and well tolerated and induced dose dependent expansion of dendritic cells in the periphery. In patients with cancer, this increase in dendritic cells can be utilized to enhance anti-tumor responses to immuno-oncology therapies.[Table: see text]

Despite the growing promise of bacterial therapy, due to the tumor targeting effect, in solid tumor treatment, most conventional therapeutic bacteria exhibit pathogenicity and insufficient therapeutic efficacy in the tumor milieu. Here we show the optical characterization of non-pathogenic natural purple photosynthetic bacteria (PPSB), which displays multifunctionality and biocompatibility in the treatment of highly active cancers. To this end, the living PPSB are applied in cancer theranostics, using bio-optical-window I and II near-infrared (NIR) light. PPSB exhibit strong NIR-I-to-NIR-II reporter fluorescence, powerful photothermal conversion, excellent reactive oxygen species generation, and contrasting photo-acoustic effects, via the energy transfer system of light harvesting nanocomplexes in PPSB membranes, making PPSB useful for highly targeted tumor elimination and precisely marking tumor location with the help of immune system. Furthermore, short-term fasting, that can evoke anticancer immunological reactions, could dramatically improve the optical efficacies of the bacterial treatment. Starving-mediated and NIR light-driven PPSB would serve as an effective “all-in-one” theranostic material for use in deep tumor treatments.

An alpha-cyclodextrin (α-CD)-based gold/DNA nanomachine was developed as a novel theranostic agent for tumor-selective diagnosis and therapy., Nanoparticles as novel theranostic agents for cancer treatment have been extensively investigated in recent years. However, the poor tumor selectivity and retention of the theranostic agents result in unsatisfactory performance of both the diagnostic and therapeutic functions. Herein, we developed an alpha-cyclodextrin (α-CD)-based gold/DNA nanomachine for tumor-selective diagnosis and therapy. The α-CDs were capped at the ends of DNA, and their release was triggered by the low pH of the tumor microenvironment, which further resulted in DNA self-assembly through complementary base pairing. The large-sized gold aggregates failed to escape from the tumor tissue, thereby realizing the goal of tumor-specific targeting and enhanced retention. Thus, the photoacoustic signal and photothermal effect are also activated, thereby achieving tumor-targeted photoacoustic imaging and photothermal therapy. In vivo results indicated that the designed gold nanomachines can serve as efficient theranostic agents for diagnosis and therapy. Moreover, we found that the α-CD caps have the ability to protect the nanoparticles from clearance and enzyme digestion, which helps the nanoparticles reach the tumor more efficiently.

Chemotherapy resistance remains a serious impediment to successful antitumor therapy around the world. However, existing chemotherapeutic approaches are difficult to cope with the notorious multidrug resistance in clinical treatment. Herein, we developed tumor-specific multiple stimuli-activated dendrimeric nanoassemblies with a metabolic blockade to completely combat both physiological barriers and cellular factors of multidrug resistance. With a sophisticated molecular and supramolecular engineering, this type of tumor-specific multiple stimuli-activated nanoassembly based on dendrimeric prodrugs can hierarchically break through the sequential physiological barriers of drug resistance, including stealthy dendritic PEGylated corona to optimize blood transportation, robust nanostructures for efficient tumor passive targeting and accumulation, enzyme-activated tumor microenvironment targeted to deepen tumor penetration and facilitate cellular uptake, cytoplasmic redox-sensitive disintegration for sufficient release of encapsulated agents, and lysosome acid-triggered nucleus delivery of antitumor drugs. In the meantime, we proposed a versatile tactic of a tumor-specific metabolism blockade for provoking several pathways (ATP restriction, apoptotic activation, and anti-apoptotic inhibition) to restrain multiple cellular factors of drug resistance. The highly efficient antitumor activity to drug-resistant MCF-7R tumor in vitro and in vivo supports this design and strongly defeats both physiological barriers and cellular factors of chemotherapy resistance. This work sets up an innovative dendrimeric nanosystem to surmount multidrug resistance, contributing to the development of a comprehensive nanoparticulate strategy for future clinical applications.

// Sameer Chopra 1,* , Jie Liu 2,* , Mehrdad Alemozaffar 1 , Peter W. Nichols 3 , Manju Aron 3 , Daniel J. Weisenberger 4 , Clayton K. Collings 1 , Sumeet Syan 1 , Brian Hu 5 , Mihir Desai 1 , Monish Aron 1 , Vinay Duddalwar 6 , Inderbir Gill 1 , Gangning Liang 1 and Kimberly D. Siegmund 2 1 Department of Urology, Norris Comprehensive Cancer Center, USC Keck School of Medicine, University of Southern California, Los Angeles, CA, USA 2 Department of Preventive Medicine, Norris Comprehensive Cancer Center, USC Keck School of Medicine, University of Southern California, Los Angeles, CA, USA 3 Department of Pathology, Norris Comprehensive Cancer Center, USC Keck School of Medicine, University of Southern California, Los Angeles, CA, USA 4 Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center, USC Keck School of Medicine, University of Southern California, Los Angeles, CA, USA 5 Department of Urology, Loma Linda University, Loma Linda, CA, USA 6 Department of Radiology, Norris Comprehensive Cancer Center, USC Keck School of Medicine, University of Southern California, Los Angeles, CA, USA * These authors have contributed equally to this work Correspondence to: Gangning Liang, email: // Kimberly D. Siegmund , email: // Keywords : small renal mass, tumor classification, DNA methylation, kidney cancer Received : July 29, 2016 Accepted : September 20, 2016 Published : September 27, 2016 Abstract Purpose: The clinical management of small renal masses (SRMs) is challenging since the current methods for distinguishing between benign masses and malignant renal cell carcinomas (RCCs) are frequently inaccurate or inconclusive. In addition, renal cancer subtypes also have different treatments and outcomes. High false negative rates increase the risk of cancer progression and indeterminate diagnoses result in unnecessary and potentially morbid surgical procedures. Experimental Design: We built a predictive classification model for kidney tumors using 697 DNA methylation profiles from six different subgroups: clear cell, papillary and chromophobe RCC, benign angiomylolipomas, oncocytomas, and normal kidney tissues. Furthermore, the DNA methylation-dependent classifier has been validated in 272 ex vivo needle biopsy samples from 100 renal masses (71% SRMs). Results: In general, the results were highly reproducible (89%, n =70) in predicting identical malignant subtypes from biopsies. Overall, 98% of adjacent-normals ( n =102) were correctly classified as normal, while 92% of tumors ( n =71) were correctly classified malignant and 86% of benign ( n =29) were correctly classified benign by this classification model. Conclusions: Overall, this study provides molecular-based support for using routine needle biopsies to determine tumor classification of SRMs and support the clinical decision-making.

Purpose The purpose of this study was to evaluate the usefulness of L-4-borono-2-18F-fluoro-phenylalanine (18F-FBPA) as a tumor-specific probe, in comparison to 18F-FDG and 11C-methionine (Met), focusing on its transport selectivity by L-type amino acid transporter 1 (LAT1), which is highly upregulated in cancers.

1991 Objectives Prostate cancer is often difficult to detect due to its small size. Recent advances in PET detector technology allows reconstruction utilizing time-of-flight post-processing, which has been shown to improve spatial resolution and image quality, particularly in obese patients. The purpose of this study is to compare SUVmax, SUVpeak, SUVaverage, and tumor volume for 18F choline PET/MRI in the evaluation of patients with newly diagnosed biopsy-proven high-grade prostate cancer compared to standard PET/MRI reconstruction without time-of-flight reconstruction. Methods From April 1, 2015 to October 30, 2015, 12 patients with biopsy proven prostate cancer were evaluated with 18F choline PET/MRI in this retrospective institutional review board-approved study. Twelve patients with newly diagnosed biopsy-proven prostate cancer underwent simultaneous 18F choline PET/MRI. Images were reviewed separately by 2 physicians with dual board certification in radiology and nuclear medicine, an abdominal imaging fellow with board certification in nuclear medicine, and a nuclear medicine fellow. SUVmax and SUVpeak were documented for dominant 18F choline avid lesions and SUVaverage was obtained of the pelvic muscle background for 1-6 minute and 7-30 minute PET acquisition data sets. TOF SUVs were compared to non-TOF SUVSs for 1-6 minute and 7-30 minute PET acquisition data sets. Subjective tumor volumes of the dominant prostate lesions were measured on 18F choline PET images with standard non-TOF and TOF post-processing for 1-6 minute and 7-30 minute PET acquisition data sets. Measurements were compared to pathologic specimens. Results TOF SUVmax and SUVpeak were significantly higher for 1-6 minute data sets (4-20% higher, p

We describe a thermodynamic-motivated, information theoretic analysis of proteomic data collected from a series of 8 glioblastoma multiforme (GBM) tumors. GBMs are considered here as prototypes of heterogeneous cancers. That heterogeneity is viewed here as manifesting in different unbalanced biological processes that are associated with thermodynamic-like constraints. The analysis yields a molecular description of a stable steady state that is common across all tumors. It also resolves molecular descriptions of unbalanced processes that are shared by several tumors, such as hyperactivated phosphoprotein signaling networks. Further, it resolves unbalanced processes that provide unique classifiers of tumor subgroups. The results of the theoretical interpretation are compared against those of statistical multivariate methods and are shown to provide a superior level of resolution for identifying unbalanced processes in GBM tumors. The identification of specific constraints for each GBM tumor suggests tumor-specific combination therapies that may reverse this imbalance.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer that is currently incurable with conventional chemotherapy. Some TNBC patients harbor either recurrent somatic gain-of-function mutations or focal amplification in genes coding for Notch receptors. Recent findings suggest that Notch signals, in the absence of Notch mutations are also important to maintain chemotherapy-resistant circulating tumor cells. Importantly, elevated Notch signaling is specifically enriched in the TNBC and is associated with worse survival in this subtype. These findings underscore the importance of Notch signaling in pathogenesis of TNBC and suggest potential new avenue for therapeutic approaches. Despite this, the target genes and oncogenic mechanisms downstream of aberrant Notch signaling are poorly understood. To create a high confidence map of the oncogenic Notch-driven regulome in TNBC, we performed an integrative, comprehensive and functional genome-wide analysis of Notch-driven targets in Notch-mutated TNBC cell lines by combining high-resolution genome-wide chromatin topology data (HiChIP), dynamic mapping of Notch transcriptional complex members and active and repressive chromatin marks (ChIP-seq), as well as evaluation of dynamic transcription (RNAseq) in presence and absence of Notch signals. One important oncogene and critical Notch target in TNBC is MYC. Our integrative analysis identified detailed map of Notch-dependent MYC regulatory network. The TNBC-specific MYC regulatory region, located 5' of MYC, is much more complex compared to other known Notch-driven malignancies and constitutes of a broad domain of active enhancers marked by H3K27ac that are not present in Notch-driven lymphoma and leukemia. Integrative analysis of ChIP-seq and HiChIP revealed multiple enhancers within this broad domain of regulatory elements that are directly bound by the members of Notch transcriptional complex, sensitive to acute changes in Notch activation and that contact MYC promoter via tumor specific chromatin loops. We showed that genetic perturbation of MYC by CRISPR/Cas9 approach was sufficient to kill TNBC cells. Since, MYC is difficult to target pharmacologically an alternative strategy is to use drugs that inhibit factors regulating MYC expression. To this end, elucidating the Notch regulatory landscape, and in particular, the mechanism by which Notch regulates MYC expression in TNBC may reveal new therapeutic strategies to precisely target MYC in Notch-dependent breast cancer. Citation Format: Petrovic J, Zhou Y, Georgakilas G, Vahedi G, Pear WS, Faryabi RB. Triple negative breast cancer-specific chromatin conformation links Notch signal to tumor-specific transcriptional program [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P2-04-07.

Tumor-specific fluorescence labeling is promising for real-time visualization of solid malignancies during surgery. There are a number of technologies to confer tumor-specific fluorescence. Antibodies have traditionally been used due to their versatility in modifications; however, their large size hampers efficient fluorophore delivery. Nanobodies are a novel class of molecules, derived from camelid heavy-chain only antibodies, that have shown promise for tumor-specific fluorescence labeling. Nanobodies are ten times smaller than standard antibodies, while maintaining antigen-binding capacity and have advantageous features, including rapidity of tumor labeling, that are reviewed in the present report. The present report reviews special considerations needed in developing nanobody probes, the status of current literature on the use of nanobody probes in fluorescence guided surgery, and potential challenges to be addressed for clinical translation.

Recently, catalytic therapy on the basis of Fenton reaction represents a very promising paradigm for potential cancer therapy, which has inspired great research interest and got fast progress. Nevertheless, it remains challenging to fabricate nanocatalytic systems that can precisely perform catalytic activity in tumor tissues without causing toxicity to adjacent normal tissues. Herein, we designed and fabricated a porous CeO2 nanorods that can selectively perform therapeutic activity in the weakly acidic tumor microenvironment via converting H2O2 into •OH to kill cancer cells, meanwhile, the normal tissues are protected by decomposing H2O2 into O2 in the physiological environment. Specifically, benefiting from the porous structure, such CeO2 nanorods could be loaded with Indocyanine green (ICG) that could both contribute itself to generate singlet oxygen (1O2) and convert near-infrared (NIR) light to hyperthermia. Once exposed to NIR light, on the one hand, the catalytic activity of CeO2 was significantly enhanced by photothermal effect of ICG for highly efficient tumor catalytic therapy; on the other hand, the generated hyperthermia and 1O2 by ICG can further synergistically enhance the therapy efficiency via directing killing cancer cells. The current study provides a proof-of-concept demonstration of tumor-specific therapy with minimal non-targeted toxicity.

200 Background: Gastric cancer was the fifth cancer world wide and the third leading cause of cancer-related death. PD-1 antibody has been approved for treatment in gastric cancer patients with positive expression of PD-L1 and microsatellite instability. MASCT-I (Multi-Antigen Stimulated Cell Therapy-I Injection) is an autologous non-engineered immune cell therapy for solid tumor, it is composed of multiple-antigen peptides(15 tumor associated antigens) loaded mature dendritic cells (DCs) and in vitro DC stimulated and proliferated effector T cells, the combination of MASCT-I and PD1 antibody could have a synergistic anti-tumor effect. Methods: This is a single center, phase I trial to explore the safety and tolerance of the combination of MASCT-I and PD-1 antibody in advanced gastric cancer (NCT03393416). 15 patients with PD-L1 expression or microsatellite instability were enrolled between April 2018, and June 2019. Camrelizumab was administered once every two weeks, DC and T cells were sequential administered once a month. Therapy continued until the disease progresses. Results: Only adverse reaction below grade 2 was observed related to MASCT-I,such as fever, fatigue etc. Abnormal liver function and reactive capillary hyperplasia may be related to Camrelizumab. Among the 15 patients, the longest treatment duration was 18.9 months, and the median progression-free survival (mPFS) was 3.93 months, among which, mPFS of patients with microsatellite instability is 10.23 months, mPFS of patients expressing PD-L1 is 2.35 months, which is slightly higher than the historical data of Camrelizumab alone in the treatment of advanced gastric cancer (8 weeks for mPFS). The patient 1002 always had high content of CD8+ cells and NK cells and the positive immune response in their bodies, which may plays a role in killing the tumor cells. Conclusions: The combination of MASCT-I and Camrelizumab in the treatment of advanced gastric cancer or gastroesophageal junction cancer is a safe treatment regimen, and its efficacy deserves further study. Clinical trial information: NCT03393416.

Cholangiocarcinoma (CCA) is an aggressive cancer with poor prognosis, where the median overall survival remains less than 1 year with standard treatment for unresectable or metastatic disease. This highlights the need for new therapeutic approaches. The fibroblast growth factor receptor (FGFR) family of transmembrane receptors, which are implicated in tumorigenesis, may represent one such target. In this article, we summarize the therapeutic rationale for targeting these receptors in CCA and the role of molecular testing to identify potential responders to treatment. We examine the current available safety and efficacy data on the several small molecule tyrosine kinase inhibitors under investigation for this disease. With a move toward identifying and targeting tumor specific mutations, FGFR inhibitors represent an exciting development which may be available for patients in the near future.