Post-treatment Biopsies Research Articles

Phase 2 trial of TRC102 (methoxyamine HCl) with temozolomide (TMZ) in patients with granulosa cell ovarian cancer.

5564 Background: TRC102 is a novel small molecule that binds to reactive aldehyde created in apurinic/apyrimidinic (AP) sites, inhibiting base excision repair (BER), which is implicated as a pathway of resistance to alkylating agents. In preclinical studies, TRC102 demonstrated synergistic anti-tumor activity when TRC102 was combined with the alkylating agent temozolomide (TMZ) and the phase 1 trial of this combination reported 4 patients with partial response, two of which were of granulosa cell ovarian cancer (GCOC) histology. Here we report the phase 2 results of the combination TRC102 and TMZ in the GCOC cohort (NCT01851369). Methods: We conducted an open-label, single center Simon 2-stage trial including patients (pts) with GCOC who received ≥ one line of therapy in the metastatic setting. TRC102 was dosed at 125 mg (100 mg for BSA < 1.6) and TMZ was dosed at 150 mg/m2 orally on day (D) 1-5 in 28-day cycle (C). Restaging CT scans every 2 Cs were evaluated by RECIST v1.1. Mandatory paired biopsies (C1D1 pretreatment and C1D4 3-4 hours after drug administration) and optional blood samples for circulating tumor cells (CTC) were collected at set time points for pharmacodynamic analyses. Results: Nine pts with GCOC (7 adult, 2 juvenile histology) were enrolled as of 12/23/2022. Median age was 53 years (range 21-79) and median prior lines of therapy was 6 (range 3-9). Most common grade (G) 1 and G2 treatment-related adverse events (trAE) were fatigue, myelosuppression, nausea, and vomiting. One pt had G3 trAE of vomiting. No toxicity-related treatment discontinuations and no treatment-related deaths were reported. The median PFS for the 8 evaluable pts was 3.7 months. One pt exited the study after one C per pt’s choice with no restaging available. Four pts had stable disease (SD) as their best response. Of those who had SD, one pt completed 26 Cs prior to progression, one pt completed 11 Cs as of data cut-off but continues on study, two pts completed 6 Cs (one pt went off study for PD and one by pt choice). MGMT analysis was performed for 3 pts, including the pt still on study after 11 Cs with SD. MGMT immunohistochemistry (IHC) of pretreatment biopsies were positive for protein expression, consistent with unmethylated MGMT status. No significant induction in the levels of the DNA damage response markers γH2AX, pNbs1, and RAD51 was detected in 5 evaluable post-treatment biopsy samples as compared to the pre-treatment timepoint. Conclusions: TRC102 combined with TMZ was well-tolerated and demonstrated durable disease control in 4 pts, which is promising in this heavily pretreated GCOC cohort. MGMT analysis suggests that unmethylated MGMT status and protein expression does not preclude response to TRC102/TMZ combination therapy. Analysis of CTCs and biopsy samples are ongoing to further elucidate possible biomarkers of response. Funded by NCI Contract No. HHSN261201500003I. Clinical trial information: NCT01851369 .

Vaginal epithelial histology before and after fractional CO2 laser in postmenopausal women: a double-blind, sham-controlled randomized trial

Postmenopausal vaginal symptoms affect over 60% of women and may substantially impact a woman's quality of life. Since 2012, fractional CO2 laser has been suggested as a treatment for this indication. Structural assessment of vaginal epithelium using microscopic biopsy examination has been used as a primary outcome measure and surrogate determinant of success of vaginal laser in previous clinical studies. This study aimed to report the effects of laser compared with sham treatment on human vaginal epithelium from postmenopausal women using microscopic examination of tissue biopsies. This single-center double-blind, sham-controlled randomized controlled trial was performed in a tertiary hospital in Sydney, Australia. A total of 49 postmenopausal women who were symptomatic of at least 1 vaginal symptom (vaginal dryness, burning, itching; dyspareunia; or dryness) were randomized to either laser or sham treatment. For this nested histologic study, participants had a pre- and post-treatment vaginal wall biopsy collected. Biopsy samples were analyzed by 3 independent specialist gynecologic pathologists and categorized as Type 1 (well-estrogenized), 2 (poorly estrogenized), or 3 (combination) mucosae. Other outcomes assessed included symptom severity (visual analog scale for symptoms including most bothersome symptom, and Vulvovaginal Symptom Questionnaire) and Vaginal Health Index. Prespecified secondary analyses of data were performed. Categorical data were analyzed using the Pearson chi-square test (or Fisher exact test if <5 in any category) or related-samples McNemar test for paired nonparametric data. Nonparametric, continuous variables were assessed using Wilcoxon signed-rank test or Mann-Whitney U test, and parametric variables with t test or 1-way analysis of variance as appropriate. All analyses were performed using SPSS software version 26.0 (IBM Corp, Armonk, NY). There was no significant difference in microscopic features of vaginal epithelium following laser or sham treatment (P=.20). Further subgroup analyses of age, menopause type, duration of reproductive life, time since menopause and BMI, still demonstrated no significant difference between laser and sham groups in histological category of vaginal epithelium. Microscopic features at pre-treatment vaginal biopsy were Type 1 in 27% (13/49). There was no significant difference in VAS score for overall vaginal symptom between those classified as Type 1 vs. Type 2/3 (VAS score overall: Type 1 vs. Type 2/3, (48.1 [95% CI 27.0, 69.2] vs. 61.5 [95% CI 49.8, 73.3]; P=.166). Data from this double-blind, sham-controlled randomized controlled trial demonstrate that fractional CO2 laser and sham treatment have a comparable histologic effect on vaginal tissue that is not significantly different. Fractional CO2 laser is not significantly different from sham treatment and should not be recommended for clinical use for postmenopausal vaginal symptoms.

148P Pattern of axillary lymph node involvement and scope of post neoadjuvant chemotherapy sentinel lymph node biopsy among breast cancer patients in developing countries

148P Pattern of axillary lymph node involvement and scope of post neoadjuvant chemotherapy sentinel lymph node biopsy among breast cancer patients in developing countries

Brief Communication on Pathologic Assessment of Persistent Stable Metastatic Lesions in Patients Treated With Anti-CTLA-4 or Anti-CTLA-4 + Anti-PD-1 Therapy.

Despite the wide use of immune checkpoint inhibition for the treatment of melanoma, the mechanisms leading to long-term stable disease are incompletely understood. Patients with metastatic melanoma who had received ipilimumab alone or ipilimumab plus nivolumab 2+years prior and attained at least 6 months of stable disease were identified. Positron emission tomography/computed tomography (PET/CT) was performed. Pretreatment and posttreatment biopsies of areas of stable disease were assessed for tumor, fibrosis, and inflammation. Seven patients underwent PET/CT and tissue biopsy. Fluorodeoxyglucose avid lesions on PET/CT ranged from no activity to an SUV of 22. In 6 patients, the residual stable lesions were composed of necrosis and fibrosis with a prominent pigment containing macrophages and no residual melanoma. In 1 patient, a nodal lesion demonstrated melanoma with active inflammation. In most patients with durable stable disease after treatment with ipilimumab or ipilimumab/nivolumab, residual lesions demonstrated predominantly necrosis and fibrosis consistent with resolving lesions. The presence of melanophages in these samples may suggest ongoing immune surveillance. One patient did demonstrate residual melanoma, indicating the need for ongoing monitoring of this patient population.

Abstract CT241: A phase 1, first-in-human (FIH) study of autologous anti-HER2 chimeric antigen receptor macrophage (CAR-M) in participants (pt) with HER2 overexpressing solid tumors

Abstract Background: Macrophages are abundant in the solid tumor microenvironment (sTME) and can exhibit both pro- and anti-tumor functions. Macrophages can be redirected by CAR expression to phagocytose cancer cells in an antigen-specific manner. CAR-M can reprogram the sTME and present neoantigens to T cells, leading to epitope spreading and anti-tumor immunity. CT-0508 is comprised of autologous monocyte-derived proinflammatory macrophages expressing an anti-HER2 CAR. Pre-clinical studies showed that CT-0508 induced targeted cancer cell phagocytosis while sparing normal cells, decreased tumor burden, prolonged survival, and was safe and effective. Notably, anti-HER2 CAR-M treatment led to activation of the sTME, with infiltration of CD8+ and CD4+ T cells, NK cells, dendritic cells, and increased activated CD8+ tumor infiltrating lymphocytes. In a pre-clinical model of advanced solid tumor resistant to PD1 blockade, mice treated with anti-HER2 CAR-M combined with a PD1 blocking antibody demonstrated improved tumor control, overall survival, and TME activation compared to either treatment alone, indicating synergy and capacity for CAR-M to sensitize solid tumors to checkpoint blockade. Methods: This Phase 1, FIH study is evaluating safety, tolerability, cell manufacturing feasibility, trafficking, TME activation, and preliminary evidence of efficacy of investigational product CT-0508 in 18 pt with locally advanced (unresectable)/metastatic solid tumors overexpressing HER2. Pt previously treated with anti-HER2 therapies are eligible. Filgrastim mobilized autologous CD14+ monocytes are collected by apheresis, followed by manufacturing and cryopreservation. Group 1 pt (n = 9; enrollment complete) received fractionated doses over Days 1, 3, and 5. Group 2 pt (n = 9) receive CT-0508 as a single infusion on D1. Additional cohorts include: CT-0508 co-administered with pembrolizumab and CT-0508 monotherapy administered intraperitoneally in pt with peritoneal predominant disease. Correlative assessments include pre- and post-treatment biopsies and blood samples for safety (immunogenicity), trafficking (qPCR, RNA in situ hybridization), CT-0508 persistence in blood and tumor, target antigen engagement, TME modulation (single cell RNA sequencing), immune response (TCR sequencing) and others. Citation Format: Yara Abdou, E. Claire Dees, Joanne Mortimer, Naoto Ueno, Melissa Johnson, Richard Maziarz, Jennifer Specht, Yuan Yuan, Paula Puhlman, Mathew Angelos, Saar Gill, Amy Ronczka, Thomas Condamine, Daniel J. Cushing, Michael Klichinsky, Debora Barton, Ramona F. Swaby, Kim Reiss Binder. A phase 1, first-in-human (FIH) study of autologous anti-HER2 chimeric antigen receptor macrophage (CAR-M) in participants (pt) with HER2 overexpressing solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT241.

Abstract LB198: A first-in-human, open-label, multicenter study of intratumoral SAR441000 (mixture of cytokine encoding mRNAs), as monotherapy and in combination with cemiplimab in patients with advanced solid tumors

Abstract Background: SAR441000 is an intratumoral (IT) therapy consisting of a mixture of mRNAs encoding the cytokines IL-12, IFN-α 2b, GM-CSF, and IL-15sushi. SAR441000 is hypothesized to induce a local and systemic immune response with reduced toxicity, and potentially enhance anti-tumor response when combined with checkpoint inhibitors. Methods: We conducted a phase 1 dose escalation and expansion study (NCT03871348) in patients with advanced/metastatic solid tumors to investigate SAR441000 as monotherapy (QW) and in combination with cemiplimab (350mg Q3W). The recommended dose (RD) of SAR44100 in combination with cemiplimab was further investigated to confirm the anti-tumor activity in patients with metastatic melanoma who had previously exhausted all standard of care options and failed treatment with anti-PD-(L)1 therapies. Serum samples and tumor biopsies were collected at baseline and post-treatment to characterize the PK/PD profile, and immune cell tumor infiltration. Tumor assessment was performed per RECIST 1.1 and iRECIST, where ≥1 non-injected lesion was considered as target lesion. Analysis was triggered after the last treated patient underwent their 2nd tumor assessment. Results: 77 patients were treated, 36 in escalation phase (21 in monotherapy, 15 in combination therapy) across eight dose levels (8µg-4000µg) and 41 in expansion phase. No dose limiting toxicities were observed. During escalation phase, 1 patient (6%) achieved confirmed complete response (iCR), 1 (6%) partial response (iPR), and 3 (20%) stable disease (iSD) as per iRECIST in combination therapy. Paired biopsies from the iPR patient showed significant increases in CD3+ and CD8+ immune cells in both injected and non-injected lesions compared to baseline. Increase of peripheral PK and PD (IFN-γ and IP-10) cytokines were observed post-treatment but no clear dose response relationship was established. Based on the overall data, 4000µg was chosen as the RD. During expansion phase, 2 (5%) melanoma patients achieved confirmed iPR and 12 (29%) achieved iSD as best overall response. Although clinically meaningful responses of injected and/or non-injected lesions were observed in 13 (32 %) patients, responses were mostly limited to loco-regional disease and no significant distant non-injected lesion response was seen. Post-treatment biopsies showed increased CD3+ and CD8+ immune cells in approximately 50% and 40% of injected and non-injected tumors, respectively. Most frequent (&amp;gt;10%) related TEAEs observed in expansion phase included injection site pain (26.8%), fatigue, diarrhea, pruritus, and asthenia (each 12.2%). Conclusion: IT with SAR441000 in combination with cemiplimab was generally well tolerated and demonstrated anti-tumor activity and immune cell infiltration in both injected and non-injected tumors. Citation Format: Oliver Bechter, Carmen Loquai, Stephane Champiat, Jean Francois Baurain, Jean-Jacques Grob, Jochen Utikal, Sylvie Rottey, Alfonso Berrocal, Jessica Hassel, Ana Arance, Miguel F. Sanmamed, Marye Boers-Sonderen, Brian Gastman, Christoffer Gebhardt, Brant Delafontaine, Ugur Sahin, Özlem Türeci, Giovanni Abbadessa, Gianfranco Di Genova, Patrick Brueck, Rahul Marpadga, Helen Lee, Céleste Lebbe. A first-in-human, open-label, multicenter study of intratumoral SAR441000 (mixture of cytokine encoding mRNAs), as monotherapy and in combination with cemiplimab in patients with advanced solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB198.

COMPLICATION AND SAFETY OF TRANSPERINEAL PROSTATE BIOPSY USING NOVEL AFFORDABLE VY PROBE (TPPB-VY) IN CLINICAL PRACTICE : A PILOT STUDY

Objective: This study was aimed to know the complication and safety of TPPB-VY in clinical practice. Material &amp; Methods: TPPB were performed in 34 patients between January 2019 and Juni 2021. Ethical approval of this study was sought from the hospital authorities (IRB number: LB.02.01/X.6.5/ 55/2020). Patients underwent systematic 10-12 core TPPB depend on the prostate volume with intravenous light sedation. The patient was included to the study if PSA ≥ 4 ng/ml and/or Digital Rectal Examination (DRE) results suggestive of prostate cancer. Procedures were performed without any antibiotic prophylaxis or an enema before the procedure and post procedure the patients just get an oral fluoroquinolones and low potent analgetics (if needed). The complication include pain level post procedure (using VAS), perineal hematoma, fever and retention post biopsy were recorded. Results: In all, 34 patients were included in the study. The result showed that perineal pain perceived by patients post procedure were 76.47% (26 out of 34 patients) but the VAS score just mild around 0-2. There were 23.53% (8 out of 34 patients) hematuria, 5.88% (2 out of 34 patients) acute urinary retention and 2.9 % (1 out of 34 patients) perineal hematoma. No fever or clinical infection was observed and 82.35% (28 out of 34 patients) patients presented with minor complications (ClavienDindo I). Only one of the patients were hospitalized due to the post-biopsy complication that need intervention. Conclusion: This is the first study TPPB using Novel Affordable VY probe. The results show clinically insignificant complication without infection. TPPB using Novel Affordable VY probe is feasible and safe.

Abstract 4353: Utilizing scRNA sequencing to understand biomarkers of response and resistance to Sacitizumab Govetican in localized TNBC

Abstract Triple-Negative Breast Cancer (TNBC) is an aggressive breast cancer subset, which lacks expression of estrogen receptors, progesterone receptors, and human epidermal growth factor receptor-2. This subset of breast cancer disproportionately affects Black and African American women and improving TNBC treatment options is vital to reducing breast cancer mortality. The novel antibody-drug conjugate, Sacitizumab Govetican (SG), which targets TROP2 at the cell surface, has shown promising clinical results from the NeoSTAR trial (NCT04230109), a phase II study evaluating neoadjuvant SG therapy in a localized TNBC setting. As part of the NeoSTAR clinical trial, we have collected and processed pre- and post- SG treatment patient samples, with the aim of understanding response to this monotherapy. Herein, we identify biomarkers of response and resistance to SG monotherapy through the use of single cell RNA sequencing of matched pre- and post-treatment patient biopsies combined with exome sequencing these patient samples. Pre-treatment core needle biopsy samples, and if applicable, post-treatment residual disease biopsies were dissociated into single cell suspensions and subjected to single cell RNA sequencing. Additionally, fixed patient tissue samples were processed accordingly for exome sequencing analyses. Overall, we analyzed over 144,000 cells from 37 total scRNA seq libraries with an average of 3800 cells per biopsy sample, demonstrating the feasibility of this method. From these analyses, we observed several cell-type differences between patients who achieved a pathological complete response (pCR) and patients who had residual disease (RD). Specifically, our data shows that tumor infiltrating lymphocytes are a potential prognostic biomarker of response to SG. Furthermore, we detected alterations in among stromal and immune cell subsets, among non-responders, indicating that these cell types maybe indicative of SG resistance. Taken together, we outline biomarkers of response to SG treatment for an improved understanding of resistance mechanisms in the neoadjuvant setting to improve TNBC outcomes among patients. Citation Format: Nicole Peiris, Simona Cristea, Mengran Zhang, Siang Koh, James Coates, Ilze Smidt, Veerle Bossuyt, Laura Spring, Aditya Bardia, Leif Ellisen. Utilizing scRNA sequencing to understand biomarkers of response and resistance to Sacitizumab Govetican in localized TNBC. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4353.

Abstract 109: Multi-modal single-cell analysis of immunotherapy-experienced cutaneous and mucosal melanoma

Abstract Immune checkpoint blockade (ICB) targeting CTLA-4 or the PD1/PD-L1 axis have become a standard of care therapy for most patients with metastatic melanoma. However, for patients with resistance to front-line ICB, especially those with BRAFWT tumors who received combination therapy, subsequent therapeutic options remain limited. Thus, understanding how ICB alters the tumor ecosystem, and thereby may create new vulnerabilities, is key to understanding effective therapies for ICB-experienced patients. Here, enabled through innovations in biospecimen research, we performed multi-modal single-cell RNA, T-cell receptor and single-cell spatial transcriptomics, coupled with whole-genome sequencing (WGS) of 42 melanoma patient biopsies. This cohort includes 7 patients with paired before- and on-/post-treatment biopsies, and spanned treatment-naïve metastatic cutaneous melanoma patients (n=10), on-ICB (aPD-1, n=7; aCTLA-4, n=1; combination therapy, n=2; total, n=10), and post-ICB (aPD-1, n=10; aCTLA-4, n=5; combination therapy, n=6; L19IL2+L16TNF, n=1; total, n=22). This cohort further includes 6 rare samples of mucosal melanoma (oral, n=4; rectal, n=2). In total, we profiled &amp;gt;320,000 transcriptomes, including &amp;gt;240,000 malignant and ~80,000 non-malignant cells, and 14,714 matched TCRs. We used a series of novel analytical approaches, including KINOMO to mitigate biases introduced by differences in gene dosage, and novel artificial intelligence frameworks, to discover differentially expressed genes and gene programs associated with treatment exposures and treatment sites. Compared to untreated specimens, there is an increase in antigen presentation (Wilcoxon rank-sum test, P=0) and reduction in lineage expression (Wilcoxon rank-sum test, P=0) in ICB-experienced tumors. Furthermore, mucosal melanomas harbor unique gene regulatory networks characterized by expression of immune evasive programs (e.g. SOX4; MAST, P=0) and exhibit lower expression of oxidative phosphorylation (Wilcoxon rank-sum test, P=0). Together, these data indicate a high degree of cellular adaption and plasticity. Through integration of single-cell, spatial and WGS, we further identify cancer clones defined by their aneuploidy patterns that confer geographically restricted immune evasion with a distinctly cold tumor microenvironment (TME), despite global T cell activation and clonal expansion. This suggest significant intralesional heterogeneity with respect to response to ICB, which may be explained by cancer cell-intrinsic factors rather than insufficient T cell responses to ICB. This work provides initial insights into spatio-temporal changes induced by ICB in cancer cells and the TME in a relatively large patient cohort. Our results indicate that functional and spatial variability exists in these tumors and may provide avenues for novel therapeutic development in ICB-experienced patients. Citation Format: Jana Biermann, Yiping Wang, Linyue Fan, Patricia Ho, Amit D. Amin, Johannes C. Melms, Somnath Tagore, Kevin Hoffer-Hawlik, Parin Shah, Neha Shaikh, Priyanka Ramaradj, Mingxuan Zhang, Antoni Ribas, Caroline Robert, Elham Azizi, Benjamin Izar. Multi-modal single-cell analysis of immunotherapy-experienced cutaneous and mucosal melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 109.

Abstract 5636: Multiplex digital spatial profiling (DSP) of proteins in the tumor microenvironment in response to GSK-3 inhibition by 9-ING-41 (elraglusib) correlates with novel immunostimulatory effects observed in vivo

Abstract Glycogen synthase kinase 3 (GSK-3) is a serine/threonine kinase with key roles in myriad biological processes such as tumor progression, and inhibition of GSK-3 using the small molecule elraglusib has shown promising preclinical antitumor activity in multiple tumor types. Our preclinical experiments showed that elraglusib treatment increased tumor cell PD-L1 expression, downregulated angiogenic and immunosuppressive signaling pathways, and increased anti-tumor immune responses in vitro and in vivo. Subsequent studies showed that several circulating factors were predictive of response to PD-1/PD-L1 blockade and GSK-3 inhibition in a murine model of colorectal cancer. To determine the translational relevance of our prior results we evaluated tumor biopsies and plasma samples from patients with refractory solid tumors of multiple tissue origins enrolled in a Phase 1 clinical trial investigating elraglusib (NCT03678883). Plasma samples were collected from patients at baseline and 24 hours post-IV administration of elraglusib and were analyzed using Luminex technology. Paired FFPE tumor biopsies from patients with colorectal or pancreatic cancer before and after treatment were selected to analyze the tumor microenvironment using NanoString GeoMx DSP technology. The region of interest (ROI) selection strategy focused on mixed tumor and immune cell segments and ROIs were segmented using panCK+ and CD45+ morphology stains. Cytokine analysis revealed that elevated baseline plasma levels of IL-1 beta and reduced levels of VEGF correlated with improved progression-free survival (PFS) and overall survival (OS). PFS was also found to be positively correlated with elevated plasma levels of immunostimulatory analytes such as Granzyme B, IFN-gamma, and IL-2 at 24 hours post-treatment with elraglusib. CD45+ tumor-infiltrating immune cells had lower expression of VISTA, PD-1, and IDO-1 inhibitory checkpoint proteins and higher expression of OX40L and B7-H3 stimulatory checkpoint proteins in post-treatment biopsies as compared to pre-treatment biopsies. Moreover, time-on-study length negatively correlated with CD39 expression in PanCK+ segments and positively correlated with CD163 expression in CD45+ segments. This ongoing study, to our knowledge, represents the first digital spatial analysis of tumor biopsies from patients treated with elraglusib. These novel circulating biomarkers of response to GSK-3 inhibition could provide significant clinical utility and the spatial proteomics data may give us insights into the immunomodulatory mechanisms of GSK-3 inhibition. Citation Format: Kelsey E. Huntington, Christoph Schorl, Shaolei Lu, Daniel Newhouse, Benedito A. Carneiro, Wafik S. El-Deiry. Multiplex digital spatial profiling (DSP) of proteins in the tumor microenvironment in response to GSK-3 inhibition by 9-ING-41 (elraglusib) correlates with novel immunostimulatory effects observed in vivo. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5636.

Abstract 6677: Preliminary immune correlatives from BCA101 trial show favorable modulation of tumor immune microenvironment

Abstract Background: BCA101 is a bispecific antibody targeting EGFR and TGF-β. TGF-β pathway activation is a hallmark of human immune-excluded tumors, and TGF-β expression is associated with resistance to anti-PD-1 blockade. Neutralization of TGF-β removes an immunosuppressive signal that drives accumulation and polarization of myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs) in solid tumors, while EGFR inhibition targets tumor cell-intrinsic oncogenic signaling. Co-targeting of EGFR and TGF-β directly impacts tumor progression while enhancing the immunogenicity of tumors. Methods: Patients with multiple solid tumor types (CRC, pancreatic, HNSCC, SqNSCLC, and others) were treated with escalating doses of either single agent BCA101 or in combination with anti-PD-1 (pembrolizumab) enrolled on NCT04429542 trial. We performed a variety of immune correlatives on pre- and on-treatment tumor biopsies, including Nanostring-based transcriptomic profiling and IHC for immunophenotypic markers, as well as multiparametric flow cytometric profiling of circulating PBMCs. Results: Our preliminary evidence suggests that neutralization of TGF-β positively alters the systemic immune state (PBMCs) and tumor immune phenotype (mRNA, IHC). Circulating HLA-DR+ monocytes were significantly increased in on-treatment PBMC samples relative to screening. Pathway analysis of on-treatment tumor biopsies revealed enhanced costimulatory signaling, cytokine and chemokine signaling, immune infiltration, and interferon signaling. Top differentially regulated genes in on-treatment biopsies included CCL21, CXCL9, CXCL11, and CXCL13, which recruit T and NK cells. HDAC11, which negative regulates type-I interferon signaling, was significantly reduced in on-treatment biopsies. Notably, two patients with EGFR-amplified squamous non-small cell lung cancer, who both progressed on first-line immunotherapy treatment, were treated with BCA101 at 1250 mg and 1500 mg qw and achieved a partial response (ongoing for 10 months at the time of the data cutoff) and a prolonged stable disease for 11 months, respectively. They exhibited increased CD8+ T cell infiltration and a reduction in TAMs following treatment. Conclusions: Increased abundance of circulating HLA-DR+ monocytes following treatment indicated polarization towards a more positive, Th1-like systemic immune state. We observed enhanced immunogenicity of tumors as assessed by a targeted IO transcriptomic analysis. The results of the pathway analysis were supported by IHC on post-treatment biopsies from a subsequent cohort showing enhanced CD8+ T cell infiltration and stable, or reduced expression of TAM marker CD163. These results indicate that neutralization of TGF-β induces a more permissive tumor immune microenvironment. Citation Format: Patrick H. Lizotte, Paul Paik, Liviu Niculescu, Seng-Lai H. Tan, David Bohr, Elham Gharakhani, Ralf Reiners, Rachel Salazar, Avanish Varshney, Shiv Ram Krishn, Pradip Nair, Cloud Paweletz. Preliminary immune correlatives from BCA101 trial show favorable modulation of tumor immune microenvironment. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6677.

Abstract 6629: Genomic and immunophenotypic landscape of acquired resistance to PD-(L)1 blockade in non-small cell lung cancer

Abstract Background: Despite immune checkpoint inhibition (ICI) has extended survival in patients (pts) with non-small cell lung cancer (NSCLC), acquired resistance to ICI develops after an initial clinical benefit. However, the mechanisms underlying the development of acquired resistance to ICI in NSCLC are largely unknown. Methods: Tumor genomic profiling, immune cell subsets assessment from standard haematoxylin/eosin (H&amp;E) stained slides using machine learning, multiplexed immunofluorescence (CD8, PD1, PD-L1, Foxp3, CK7), and HLA-I immunohistochemistry (IHC) were performed on matched pre and post ICI tumor biopsies from pts with NSCLC treated with ICI at the Dana-Farber Cancer Institute who developed acquired resistance to ICI. Results: Among 1823 pts with advanced NSCLC who received ICI, 80 developed acquired resistance to treatment and had matched pre- and post-ICI tissue samples for correlative analysis. Putative resistance mutations were identified in 27.3% of cases. The most common acquired loss-of-function mutations included STK11 (9.1%), B2M (6.5%), NF1/2 (5.2%), SMARCA4 (5.2%), APC (3.9%), KEAP1 (2.6%), and JAK1 (2.6%). In addition, acquired activating mutations were found in PI3KCA (3.9%) and SOS1 (1.3%). In examining copy number variations (CNVs), we identified acquired CNVs in 49.3% (38/77) of ICI-resistant samples, including heterozygous loss of B2M (23.4%), STK11 (16.8%), SMARCA4 (16.9%), KEAP1 (15.6%), PTEN (10.4%), CDKN2A/B (9.1%) and PD-L1/PD-L2 genes (9.1%), among others. Importantly, we also noted acquired homozygous deletions in CDKN2A/B (9.1%), PD-L1/PD-L2 (1.3%) and JAK2 (1.3%), and amplification in MDM2 (5.2%), KRAS (3.9%) and MYC (2.6%), at the time of resistance to ICI. Among 16 cases with pre- and post-ICI H&amp;E stained slides, digital pathology analysis showed a significant decrease in the density of intratumoral lymphocytes in ICI-resistant tumor samples compared to pre-ICI samples (88 vs 36 cells/mm2, P=0.03). Consistently, among 6 pts with pre- and post-ICI multiplexed immunofluorescence we observed a significant decrease in CD8+ PD1+ T cells in ICI-resistant vs pre-ICI tumor samples (24 vs 10 cells/mm2, P=0.01). Among pts with tissue available for HLA-I IHC (N=9), we found a significant decrease in HLA-I expression by H-score at the time of acquired resistance (300 vs 260, P=0.03). In control cohorts of pts with pre- and post-chemotherapy (CT) (N=32) or targeted therapies (TT) (N=89) tumor genomic profiling no acquired mutations in STK11, KEAP1, B2M, JAK1, APC, SOS1 were detected. Similarly, intervening chemotherapy and targeted therapies were not associated with changes in lymphocytes density and HLA-I expression. Conclusion: Mechanisms of acquired resistance to ICI are heterogenous, including both genomic and immunophenotypic factors. New therapeutic strategies are required to delay and overcome ICI resistance in pts with NSCLC. Citation Format: Biagio Ricciuti, Giuseppe Lamberti, Sreekar Puchala, Navin Mahadevan, Joao Alessi, Xinan Wang, Yvonne Li, Federica Pecci, Alessandro Di Federico, Malini Gandhi, Adriana Barrichello, Victor Vaz, Andy Pangilinan, Danielle Haradon, Elinton Lee, Hersh Gupta, Mizuki Nishino, Andrew Cherniack, Scott Rodig, Lynette Sholl, Renato Umeton, MArk M. Awad. Genomic and immunophenotypic landscape of acquired resistance to PD-(L)1 blockade in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6629.

PD15-03 RADIATION DOSE REQUIRED TO CONTROL PROSTATE CANCER AS DETERMINED BY POST-TREATMENT BIOPSY: LONG TERM RESULTS

You have accessJournal of UrologyCME1 Apr 2023PD15-03 RADIATION DOSE REQUIRED TO CONTROL PROSTATE CANCER AS DETERMINED BY POST-TREATMENT BIOPSY: LONG TERM RESULTS Nelson Stone, Rendi Shue, Barry Rosenstein, and Richard Stock Nelson StoneNelson Stone More articles by this author , Rendi ShueRendi Shue More articles by this author , Barry RosensteinBarry Rosenstein More articles by this author , and Richard StockRichard Stock More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003262.03AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Radiation therapy for prostate cancer is typically prescribed at a standard dose, 79-81 Gy for external beam irradiation (EBRT) and 145 Gy for iodine-125 brachytherapy (BT). Unlike prostatectomy where the gland is removed at the time of treatment successful radiation treatment relies on the death of all cancer cells in situ. However, the dose requirement for different Gleason grade groups or NCCN risk groups may also differ. We investigated the relationship between the biological effective dose (BED) and local control as determined by post-treatment biopsy (PB) in a large cohort of men who had BT +EBRT. METHODS: 545 men who had BT were followed a median of 11 years (range 2-26) and had PB a mean of 3.2 years (2-17) post-treatment. All PB were performed by one investigator (NNS) and routinely performed at 2 years or later for a rising PSA. Median age, PSA and BED were 66 years (41-85), 7.0 ng/ml (1-189) and 185.5 Gy (15-282). 295 (54.1%) received hormone therapy (HT) for a median of 6 months. Radiation doses were converted post-implant to the BED using an ꭤ/β of 2. Associations with grade, stage, PSA, NCCN status, HT use, BED were compared to the PB results and prostate cancer survival (PCS) using ANOVA, chi-square, linear regression, Kaplan Meier estimates with comparisons by log rank and Cox proportion hazards. RESULTS: 34 (6.2%) of the 545 men had a positive PB which occurred a mean of 5.9 years (range 2-17) after all treatment. BED (p<0.001), stage (p=0.004), PSA (p=0.038) and HT use (p=0.019) were associated with a positive PB but only BED (p<0.001) remained significant on regression analysis. Dichotomizing BED to <140, 160, 180, 200, 220 or greater yielded optimal local control (positive biopsy rate 2% or less) at 180 Gy for all 3 NCCN risk groups (Table). Men with a positive PB had a mean BED of 136.8 vs. 181.3 Gy for negative PB (p<0.001). At <180 Gy 24/229 (10.5%) had local failure compared to 5/392 (1.7%) for higher doses (p<0.001, OR 6.7, 95% CI 2.5-17.9). PCS at 15 and 20 years for negative PB was 94.5 and 94.5% vs. 74.4 and 46.5% for a positive one (p<0.001). PCS was associated with BED (p<0.001), PSA (p=0.028) and stage (p=0.018). CONCLUSIONS: This long-term investigation defined the radiation dose necessary to eradicate prostate cancer and should be considered as the minimal requirement when prescribing and delivering this treatment option. Source of Funding: None © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e419 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Nelson Stone More articles by this author Rendi Shue More articles by this author Barry Rosenstein More articles by this author Richard Stock More articles by this author Expand All Advertisement PDF downloadLoading ...

258 Analysis of the Hepatic Microenvironment Before and After Direct-Acting Antiviral (DAA) Therapy for Viral Hepatitis C

OBJECTIVES/GOALS: The effect of the Direct-Acting Antiviral (DAA) on hepatic histopathological features from patients treated for HCV has not been thoroughly evaluated. The goals of this retrospective study were to determines differences between the liver biopsies collected before and after DAA treatment and correlated the histopathology with clinical outcome. METHODS/STUDY POPULATION: Spectral imaging was used to evaluate differences in intrahepatic macrophage (CD68, CD14, CD16, MAC 387, and CD163) and T cell (CD3, CD4, CD8, CD45, and FoxP3) phenotypes in paired liver biopsies collected from the same patient before (n=10) and after (n=10) achieving SVR (Figure 1). Imaging analysis and machine learning algorithms were used to evaluate changes in these key immune cells. We also compared differential gene expression of over 700 genes using RNA isolated from liver biopsies with NanoString. RESULTS/ANTICIPATED RESULTS: Multispectral imaging analysis showed a significant increase of proinflammatory/M1-like (e.g., CD14+) and anti-inflammatory/M2-like macrophage (e.g., CD163+) phenotypes in pre-treatment versus post-treatment biopsies, respectively. Gene expression analysis revealed enrichment of inflammatory (HLA-B, STAT1, CXCL10) and interferon induced-antiviral (ISG15, OAS3, MX1 and IFIT1) genes in the pre-treatment vs the post-treatment group. Cell deconvolution analysis also showed a significant increase of M1-like macrophages in the pre-treatment group when compared to the post-treatment group or controls. Upregulation of genes associated with cell proliferation and differentiation (c-KIT and Fos) was observed in the post-treatment biopsies of patients with persistent inflammatory infiltrates. DISCUSSION/SIGNIFICANCE: Protein and gene expression profiles observed in patients before DAA therapy showed a strong macrophage-mediated inflammatory response against HCV infection in the liver that shifted significantly to a tissue remodeling microenvironment after treatment.

How I treat refractory/relapsed diffuse large B‐cell lymphomas with CD19‐directed chimeric antigen receptor T cells

Chimeric antigen receptor (CAR) T cells targeting CD19 represent a promising salvage immunotherapy for relapsed/refractory diffuse large B-cell lymphoma (R/R DLBCL), offering ~40% of long-term responses. In everyday clinical practice, haematologists involved in CAR T cell treatment of patients with R/R DLBCL have to deal with diagnostically complex cases and difficult therapeutic choices. The availability of novel immunotherapeutic agents for R/R DLBCL and recent advances in understanding CAR T-cell failure mechanisms demand a rational approach to identify the best choice for bridging therapy and managing post-CAR T-cell therapy relapses. Moreover, positron emission tomography/computerised tomography may result in false-positive interpretation, highlighting the importance of post-treatment biopsy. In this review, we discuss all above issues, presenting four instructive cases, with the aim to provide criteria and new perspectives for CAR T-cell treatment of DLBCL.

Phase 2 Trial of Stereotactic Ablative Radiotherapy for Patients with Primary Renal Cancer

BackgroundMost renal cell carcinomas (RCCs) are localized and managed by active surveillance, surgery, or minimally invasive techniques. Stereotactic ablative radiation (SAbR) may provide an innovative non-invasive alternative although prospective data are limited. ObjectiveTo investigate whether SAbR is effective in the management of primary RCCs. Design, setting, and participantsPatients with biopsy-confirmed radiographically enlarging primary RCC (≤5 cm) were enrolled. SAbR was delivered in either three (12 Gy) or five (8 Gy) fractions. Outcome measurements and statistical analysisThe primary endpoint was local control (LC) defined as a reduction in tumor growth rate (compared with a benchmark of 4 mm/yr on active surveillance) and pathologic evidence of tumor response at 1 yr. Secondary endpoints included LC by the Response Evaluation Criteria in Solid Tumors (RECIST 1.1), safety, and preservation of kidney function. Exploratory tumor cell–enriched spatial protein and gene expression analysis were conducted on pre- and post-treatment biopsy samples. Results and limitationsTarget accrual was reached with the enrollment of 16 ethnically diverse patients. Radiographic LC at 1 yr was observed in 94% of patients (15/16; 95% confidence interval: 70, 100), and this was accompanied by pathologic evidence of tumor response (hyalinization, necrosis, and reduced tumor cellularity) in all patients. By RECIST, 100% of the sites remained without progression at 1 yr. The median pretreatment growth rate was 0.8 cm/yr (interquartile range [IQR]: 0.3, 1.4), and the median post-treatment growth rate was 0.0 cm/yr (IQR: –0.4, 0.1, p < 0.002). Tumor cell viability decreased from 4.6% to 0.7% at 1 yr (p = 0.004). With a median follow-up of 36 mo for censored patients, the disease control rate was 94%. SAbR was well tolerated with no grade ≥2 (acute or late) toxicities. The average glomerular filtration rate declined from a baseline of 65.6 to 55.4 ml/min at 1 yr (p = 0.003). Spatial protein and gene expression analyses were consistent with the induction of cellular senescence by radiation. ConclusionsThis clinical trial adds to the growing body of evidence suggesting that SAbR is effective for primary RCC supporting its evaluation in comparative phase 3 clinical trials. Patient summaryIn this clinical trial, we investigated a noninvasive treatment option of stereotactic radiation therapy for the treatment of primary kidney cancer and found that it was safe and effective.

Initiation of renin-angiotensin system inhibitors and first complete remission in patients with primary nephrotic syndrome: a nationwide cohort study.

Evidence on renin-angiotensin system inhibitors (RASis) effect in reducing urinary protein levels in patients with nephrotic syndrome is insufficient. We determined whether RASis can induce complete remission (CR) in patients on immunosuppressive therapy. This cohort study included 84 adults (median age, 65years; males, 57%) with primary nephrotic syndrome (excluding minimal change disease) not receiving RASis during enrollment in the Japanese Nephrotic Syndrome Cohort Study from January 2009 to December 2010, and were followed up for 5years. Exposure and outcome were RASi initiation and first CR, respectively. Marginal structural models and Poisson regression were used to account for time-varying covariates and estimate causal effects of RASis on CR. Overall, 51 (61%), 73 (87%), and 55 (66%) patients had membranous nephropathy, were prescribed immunosuppressive agents at baseline (1-month post-renal biopsy and/or at start of immunosuppressive therapy), and were prescribed RASis during the study period, respectively. Sixty-five patients experienced firstCR (incidence rate, 5.05/100 person-months). RASi use was associated with a higher (adjusted incidence rate ratio [aIRR] 2.27, 95% confidence interval [CI] 1.06-4.84), and lower (aIRR: 0.17, 95% CI 0.04-0.68) firstCR in patients with membranous nephropathy and other pathologies, respectively. RASis are beneficial as adjuvant therapy for inducing remission in patients with membranous nephropathy.

Trial in progress: Natural killer (NK) cells with TGFβ receptor I inhibitor vactosertib and IL-2 in patients with metastatic colorectal cancer or hematologic malignancies.

TPS273 Background: Natural killer (NK) cell therapy is emerging as a promising therapy for patients (pts) with solid tumors and hematologic malignancies with no reported toxicity associated with the cell therapy product itself. NK cell trials in solid tumors are limited by immune inhibitory tumor microenvironments that prevent trafficking and activity of donor NK cells, which is largely governed by TGFβ signaling. We recently developed a novel platform for ex vivo expansion of healthy donor NK cells. We previously showed these cells to be safe in a dose-escalation study when given without cytokine support, with early evidence of response in pts with metastatic colorectal cancer (CRC) and in pts with hematologic malignancies. IL-2 has been well-established as an important activating cytokine supporting NK cells in humans, but can also induce regulatory T cells, which further suppress NK cells by producing TGFβ. We seek to improve upon the early activity we previously observed with NK cells alone by administering concurrently with IL-2 and by limiting the inhibitory tumor microenvironment and regulatory T cells with TGFβ receptor 1 inhibitor vactosertib. Methods: This phase Ib open-label study evaluates the safety and persistence (primary objectives), and the clinical and biological activity and trafficking (secondary objectives) of our novel NK cell therapy in combination with IL-2 (aldesleukin) and TGFβ receptor 1 inhibitor vactosertib. Up to 12 pts with metastatic CRC or relapsed/refractory hematologic malignancies will be enrolled. Pts will undergo low-dose lymphodepletion with fludarabine and cyclophosphamide followed by 2 infusions of NK cells, administered 14 days apart. Concurrently, pts will receive aldesleukin subcutaneously 3 times weekly and vactosertib orally 5 consecutive days/week x4 weeks total. Pts will undergo pre- and post-treatment biopsies for quantification of NK cell trafficking and exploration of immune cell profiling changes in the tumor microenvironment. The trial is currently open to enrollment, with 1 pt accrued at time of submission. Clinical trial information: NCT05400122 .

Role of pre and post incisional biopsy levels of Serum Cytokeratin-19 for detection of the dissemination of oral squamous cell carcinoma.

Objective: To evaluate the diagnostic efficacy of cytokeratin 19 for the presence and spread of oral squamous cell carcinoma following incisional biopsy. Study Design: Descriptive Cross-sectional. Setting: Isra Dental College Hospital, Isra University Hyderabad. Period: November 2020 to October 2021. Material &amp; Methods: Consenting patients of either gender, age 20-70 years, with clinical suspicion of OSCC were included in the study. Patients with a previous history of chemotherapy and radiotherapy, those having history of previous biopsy, recurrence of carcinoma were excluded. After detailed examination, blood samples of all patients were collected before and after incisional biopsy and sent to the laboratory for further evaluation for CK-19 level. Results: Of total 60 patients, majority (86.6%) were male. Over half (53.3%) of the participants belongs to age-group 40-59 years. Before incisional biopsy, all participating OSCC patients were negative for the Cytokeratin RT-PCR assay. While RT-PCR (CK-19) was detected positive for 12 (20%) of patients within 15 minutes of biopsy whereas, remaining 48 (80%) showed negativity for CK-19 assay. Statistically significant difference (p&lt;0.05) was observed for the post incisional biopsy CK-19 +ve and –ve patients. Male patients and patients well differentiated OSCC tumors had positive post incisional biopsy PCR compared with their counterparts (p&lt;0.05). Conclusion: Incisional biopsy promotes the propagation of cancerous cells and increases the risk of developing metastatic lesions. Additionally, RT-PCR is a reliable tool for the detection of CK-19 transcripts in the blood.

P060 Restoration of occludin and claudin-1 expression in patients with Crohn’s disease receiving anti-TNF treatment.

Abstract Background Occludin and claudin-1 are integral components of tight junctions (TJs) that contribute to intestinal barrier integrity. The aim of this study was to prospectively evaluate the expression of occludin and claudin-1 in the intestinal mucosa of patients with Crohn’s disease (CD) undergoing treatment with tumor necrosis factor (TNF) inhibitors. Methods Clinical assessment, laboratory testing, and colonoscopy were performed on all participants prior to receiving anti-TNF therapy. Colonic or ileal biopsies were obtained from the affected intestinal areas. A follow-up exam and colonoscopy were performed at least 6 months after the start of anti-TNF therapy, and paired biopsies were obtained from the initially affected segments. The expression of the TJ proteins was evaluated by immunohistochemistry (IHC). Results In this prospective observational study, 21 consecutive patients with active CD [median age 32.5 years (IQR: 18.5-51), 66.7% male] and 10 healthy controls [36 years (IQR: 29-58), 60% male] were enrolled. Patients with CD received treatment with TNF inhibitors (infliximab, adalimumab, and certolizumab: 61.9%, 33.3%, and 4.8%, respectively), according to current guidelines, and a follow-up examination was conducted after a median period of 25 months (IQR: 9.5-66). Before treatment administration, patients with active CD exhibited an increased expression of both occludin [2 (IQR: 1.75-2.75) vs. 1.5 (IQR: 0-1.75); p = 0.032] and claudin-1 [2.5 (IQR: 2-3) vs. 1 (IQR: 0.75-2); p = 0.01] in comparison to healthy controls. In control mucosa, IHC staining for occludin and claudin-1 was weak to moderate, cytoplasmic, and membranous, confined to the surface epithelium, while staining in intestinal crypts was negative. On the other hand, an enhanced expression of both proteins in the enteric crypts and surface epithelium was demonstrated in CD patients upon initial examination. Treatment with TNF inhibitors resulted in a significant reduction of claudin-1 expression (p = 0.018) as well as a notable decline of occludin expression (p = 0.08). Occludin and claudin-1 immunostaining was restricted to the surface epithelium in post-treatment biopsies. In addition, the activity of the disease, as estimated by the Crohn’s Disease Activity Index (r = 0.476; p = 0.014) and C-reactive protein (r = 0.411; p = 0.039), was positively correlated with the expression of claudin-1 in the mucosa. Conclusion Active CD is plagued with a disruption of the expression and distribution of claudin-1 and occludin throughout the affected intestinal mucosa. Successful treatment with biological agents induces the restoration of intestinal barrier function, which is associated with an improvement in the patients' clinical condition.