Biology subjects

Alam, I. S.

Publications and source records attributed to Alam, I. S..

5 recordsLinked to original sources

bioRxiv2024

In vivo imaging of T-cell coregulator B7-H4 reveals protumor macrophage status in prostate cancer

BackgroundB7-H4 is a cell surface ligand overexpressed by tumors to inhibit T cell functions and evade the immune system. B7-H4 is minimally expressed in normal tissues but is highly expressed by various cancer cells and tumor-associated macrophages (TAM). Despite its importance as an immune checkpoint inhibitor, no imaging techniques specifically targeting B7-H4 have been established. To close this gap, we sought to assess the ability of a monoclonal antibody (mAb) based immunoPET radiotracer to visualize B7-H4 in human and murine prostate cancer models. MethodsAnti-B7-H4 mAb clone 2H9 was functionally characterized for binding to the human and mouse B7-H4 protein. The antibody was conjugated with chelator p-SCN-Bn-Deferoxamine (DFO) and labeled with radioisotope Zirconium-89 (89Zr) to obtain immunoPET tracer 89Zr-2H9-mAb. The biolayer interferometry method was used to test the binding kinetics of DFO-2H9-mAb compared to that of parental 2H9 mAb. A group of six athymic nude mice with human DU145 prostate tumor xenograft underwent MicroPET imaging after tail vein injection of [~]150{micro}Ci 89Zr-2H9-mAb or non-binding 89Zr-Isotype-mAb. Next, immunocompetent C57BL/6J mice with TRAMP-C2 tumors each were injected with either PBS (n=8), cold 2H9 mAb (10mg/kg) to block B7-H4 (n=6), or chlodronate liposome (15mg/kg) to cause total macrophage depletion (n=6), followed by 89Zr-2H9-mAb MicroPET imaging. An ex vivo biodistribution assay was performed after 144 hr post radiotracer injection. Tumor radiotracer binding, quantified as a percentage injected dose per gram (%ID/g), was compared between different experimental groups using two-way ANOVA with Bonferroni or Tukey corrections. ResultsImmunoconjugation yielded a 2.59 {+/-} 0.08 chelator-to-antibody ratio, and the binding of DFO conjugated 2H9-mAb was similar to that of parental 2H9 mAb, with unaffected affinity in targeting B7-H4 protein moiety. The radiochemical purity of 89Zr-2H9-mAb tracer was yielded >95% with an average specific activity of 5{micro}Ci/{micro}g antibody. DU145 tumor xenografts demonstrated significantly stronger radiotracer binding at 24, 48, 72, 96, and 120 hr than the non-binding isotype control group. In TRAMP-C2 tumor xenografts, the radiotracer binding in B7-H4 blocked tumors was significantly lower than in the non-blocked PBS-injected group. Macrophage depletion resulted in a significant decrease in tumor binding compared to the control group. 89Zr-2H9-mAb could efficiently distinguish tumors with high sensitivity, showing a high correlation between PET imaging and bio-distribution. More importantly, the immunohistochemistry of the harvested tumor revealed no significant difference between the three groups, as discernible through in vivo PET imaging. ConclusionThis study highlights the potential of B7-H4 immunoPET imaging for monitoring immunotherapy response. With the emerging potential of B7-H4 blocking as an immunotherapeutic, immunoPET imaging could be readily expanded to patient stratification and therapy monitoring. B7-H4 imaging could augment our understanding of B7-H4 dynamics in response to various therapeutic interventions in clinical trials. The new B7-H4 immunoPET probe is, in principle, clinically translatable.

immunology↗

bioRxiv2022

Highly Excretable Gold Supraclusters for Translatable In Vivo Raman Imaging of Tumors

Raman spectroscopy provides excellent specificity for in vivo preclinical imaging through a readout of fingerprint-like spectra. To achieve sufficient sensitivity for in vivo Raman imaging, metallic gold nanoparticles larger than 10 nm were employed to amplify Raman signals via surface-enhanced Raman scattering (SERS). However, the inability to excrete such large gold nanoparticles has restricted the translation of Raman imaging. Here we present Raman-active metallic gold supraclusters that are biodegradable and excretable as nanoclusters. Although the small size of the gold nanocluster building blocks compromises the electromagnetic field enhancement effect, the supraclusters exhibit bright and prominent Raman scattering comparable to that of large gold nanoparticle-based SERS nanotags due to high loading of NIR-resonant Raman dyes and much suppressed fluorescence background by metallic supraclusters. The bright Raman scattering of the supraclusters was pH-responsive, and we successfully performed in vivo Raman imaging of acidic tumors in mice. Furthermore, in contrast to large gold nanoparticles that remain in the liver and spleen, the supraclusters dissociated into small nanoclusters, and 73% of the administered dose to mice was excreted over 4 months. The highly excretable Raman supraclusters demonstrated here offer great potential for clinical applications of in vivo Raman imaging by replacing non-excretable large gold nanoparticles.

bioengineering↗

bioRxiv2021

Multi-parameter optical imaging of immune cell activity in chimeric antigen receptor T-cell and checkpoint blockade therapies

Longitudinal multimodal imaging presents unique opportunities for noninvasive surveillance and prediction of treatment response to cancer immunotherapy. In this work we first designed a novel granzyme B activated self-assembly small molecule, G-SNAT, for quantitative assessment of cytotoxic T lymphocyte mediated cancer cell killing in vivo. In lymphoma tumor bearing mice, the retention of cyanine 5 labeled G-SNAT-Cy5 was shown to be highly correlated to CAR T-cell mediated granzyme B release and tumor eradication. In colorectal tumor-bearing transgenic mice, expressing firefly luciferase in hematopoietic cells, and which received combination treatment of anti-PD-1 and anti-CTLA-4, longitudinal bioluminescence and fluorescence imaging revealed the dynamics of immune cell expansion, trafficking, tumor infiltration, and cytotoxic activity which predicted therapeutic outcome before tumor shrinkage was evident. These results support further development of G-SNAT for imaging early immune response to checkpoint blockade and CAR T-cell therapy in patients and highlight the utility of multimodality imaging for improved mechanistic insights into cancer immunotherapy.

immunology↗

bioRxiv2021

Engineering genetically-encoded synthetic biomarkers for breath-based cancer detection

Breath analysis holds great promise for rapid, noninvasive early cancer detection; however, clinical implementation is impeded by limited signal from nascent tumors and high background expression by non-malignant tissues. To address this issue, we developed a novel breath-based reporter system for early cancer detection using D-limonene, a volatile organic compound (VOC) from citrus fruit that is not produced in humans, in order to minimize background signal and maximize sensitivity and specificity for cancer detection. We metabolically engineered HeLa human cervical cancer cells to express limonene at levels detectable by mass spectrometry by introducing a single plant gene encoding limonene synthase. To improve limonene production and detection sensitivity twofold, we genetically co-expressed a modified form of a key enzyme in the cholesterol biosynthesis pathway. In a HeLa xenograft tumor mouse model, limonene is a sensitive and specific volatile reporter of tumor presence and growth, permitting detection of tumors as small as 5 mm. Moreover, tumor detection in mice improves proportionally with breath sampling time. By continuously collecting VOCs for 10 hours, we improve sensitivity for cancer detection 100-fold over static headspace sampling methods. Whole-body physiologically-based pharmacokinetic (PBPK) modeling and simulation of tumor-derived limonene predicts detection of tumors as small as 7 mm in humans, equivalent to the detection limit of clinical imaging modalities, such as PET, yet far more economical. Significance StatementWe developed a breath-based reporter system using the plant terpene, D-limonene - a volatile secondary metabolite that gives citrus fruit its characteristic scent but is not produced in human tissues - as a biomarker for early cancer detection. Results from this study could pave the way for in vivo gene delivery and tumor-specific expression of exogenous volatile cancer reporters with broad applicability to the early diagnosis of a wide variety of cancers.

bioengineering↗

bioRxiv2019

Metabolic Architecture of the Deep Ocean Microbiome

The deep sea, the largest compartment of the ocean, is an essential component of the Earth system, but the functional exploration of its microbial communities lags far behind that of other marine realms. Here we analyze 58 bathypelagic microbial metagenomes from the Atlantic, Indian, and Pacific Oceans in an unprecedented sampling effort from the Malaspina Global Expedition, to resolve the metabolic architecture of the deep ocean microbiome. The Malaspina Deep-Sea Gene Collection, 71% of which consists of novel genes, reveals a strong dichotomy between the functional traits of free-living and particle-attached microorganisms, and shows relatively patchy composition challenging the paradigm of a uniform dark ocean ecosystem. Metagenome Assembled Genomes uncovered 11 potential new phyla, establishing references for deep ocean microbial taxa, and revealed mixotrophy to be a widespread trophic strategy in the deep ocean. These results expand our understanding of the functional diversity, metabolic versatility, and carbon cycling in the largest ecosystem on Earth.\n\nOne Sentence SummaryA whole community genomic survey of the deep microbiome sheds light on the microbial and functional diversity of the dark ocean.

microbiology↗