CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Patent Application Serial No. 63/398,069, filed on August 15, 2022. The disclosure of the prior application is considered part of, and is incorporated by reference in, the disclosure of this application.

TECHNICAL FIELD

This document relates to methods and materials for assessing and/or using antisenescence treatments (e.g., admini strati on(s) of one or more senotherapeutic agents) within a mammal (e.g., a human). In some cases, the methods and materials provided herein can be used to determine and/or classify the efficacy of an anti-senescence treatment in a mammal (e.g., a human). For example, the level of pregnancy-associated plasma protein-A (PAPP-A) polypeptides in a sample obtained from a mammal (e.g., a human) having been administered an anti-senescence treatment (e.g., admini stration(s) of one or more senotherapeutic agents) can be used to determine and/or classify the efficacy of the anti-senescence treatment in treating the mammal.

BACKGROUND INFORMATION

PAPP-A is a secreted zinc metalloprotease. The major function of PAPP-A polypeptides is to enhance insulin-like growth factor (IGF) signalling through their ability to degrade inhibitory IGF binding proteins (IGFBPs) such as IGFBP2, IGFBP-5, and IGFBP-4. PAPP-A polypeptides can modulate IGF bioavailability in the paracrine and autocrine pathways independently of systemic levels of IGF 1. The PAPP-A-IGF pathway plays a role in cell growth, differentiation, and transformation.

SUMMARY

This document provides methods and materials for assessing the efficacy of and/or using an anti-senescence treatment (e.g., admini stration(s) of one or more senotherapeutic agents). In some cases, the methods and materials provided herein can be used to determine the efficacy of an anti-senescence treatment in a mammal (e.g., a human). For example, the level of PAPP-A polypeptides in a sample (e.g., a blood sample, a urine sample, or a saliva sample) obtained from a mammal (e.g., a human) having been administered one or more senotherapeutic agents can be used to determine and/or classify the efficacy of the one or more senotherapeutic agents. In some cases, this document provides methods and materials for treating a mammal (e.g., a human) having a disease or disorder characterized by the presence of senescent cells or an elevated number of senescent cells. For example, an antisenescence treatment (e.g., one or more senotherapeutic agents) can be administered to a mammal having a disease or disorder characterized by the presence of senescent cells or an elevated number of senescent cells, and the level of PAPP-A polypeptides within a sample obtained from the mammal can be assessed to determine and/or classify whether or not antisenescence treatment effectively reduced the number of senescent cells within the mammal. In such cases, a reduced level of PAPP- A polypeptides within the mammal can indicate that the administered anti-senescence treatment effectively reduced the number of senescent cells within that mammal, while a lack of a reduced level of PAPP-A polypeptides within the mammal can indicate that the administered anti-senescence treatment did not effectively reduce the number of senescent cells within that mammal.

As demonstrated herein, senescent cells can secrete PAPP-A polypeptides and/or can promote the expression of PAPP -A polypeptides from other cells. As also described herein, treating a mammal with an anti-senescence treatment (e.g., administering one or more senotherapeutic agents to a mammal) can reduce the number of senescent cells within the mammal such that the level of PAPP-A polypeptides within the mammal is reduced. In some cases, when the level of PAPP- A polypeptides within the mammal is reduced following a particular anti-senescence treatment, then that particular anti-senescence treatment can be continued within that mammal. In some cases, when the level of PAPP -A polypeptides within the mammal is not reduced following a particular anti-senescence treatment, then that particular anti-senescence treatment can be discontinued within that mammal. In such a case, the mammal can be treated with an alternative anti-senescence treatment. For example, the same agents can be administered to the mammal but at an increased dosing level (e.g., higher amount of agent(s) or more frequent administration of the same amount of agent(s)) or a different agent or set of agents can be administered to the mammal. As described herein, the level of PAPP- A polypeptides within a mammal can be monitored during an anti-senescence treatment to assess effectiveness of the anti-senescence treatment and/or to make adjustments to an anti-senescence treatment when it is determined to be not effective.

In general, one aspect of this document features methods for assessing efficacy of an anti-senescence treatment. The methods can include, consist of, or consist essentially of, (a) determining whether or not a mammal has a reduced level of PAPP- A polypeptides following administration of one or more senotherapeutic agents to the mammal as compared to the level of PAPP-A polypeptides within the mammal prior to the administration of the one or more senotherapeutic agents; (b) classifying the anti-senescence treatment as being effective in the mammal if the mammal has the reduced level; and (c) classifying the anti-senescence treatment as not being effective in the mammal if the mammal does not have the reduced level. The mammal can be a human. The determining step can include comparing the level of PAPP-A polypeptides within a first sample obtained from the mammal prior to the administration of the one or more senotherapeutic agents to the level of PAPP- A polypeptides within a second sample obtained from the mammal after the administration of the one or more senotherapeutic agents. The first and second samples can be blood samples. The first and second samples can be urine samples. The first and second samples can be saliva samples. The one or more senotherapeutic agents can be dasatinib, quercetin, navitoclax, Al 331852, Al 155463, fisetin, luteolin, geldanamycin, tanespimycin, alvespimycin, piperlongumine, panobinostat, FOXO4-related peptides, nutlin-3a, ruxolitinib, metformin, or rapamycin. The method can include determining that the mammal has the reduced level and classifying the anti-senescence treatment as being effective in the mammal. The method can include administering the anti-senescence treatment to the mammal after the determining step. The method can include determining that the mammal does not have the reduced level and classifying the anti-senescence treatment as not being effective in the mammal. The method can include administering a revised anti-senescence treatment that can be different from the anti-senescence treatment administered to the mammal after the determining step. The revised anti-senescence treatment can include administering a higher amount of the one or more senotherapeutic agents of the anti- senescence treatment. The revised anti-senescence treatment can include administering one or more senotherapeutic agents different from the one or more senotherapeutic agents of the anti- senescence treatment. The method can include administering the anti-senescence treatment to the mammal prior to the determining step.

In another aspect, this document features methods for treating a mammal. The methods can include, consist of, or consist essentially of, (a) determining whether or not a mammal has a reduced level of PAPP- A polypeptides following a previous administration of one or more senotherapeutic agents to the mammal as compared to the level of PAPP -A polypeptides within the mammal prior to the previous administration; (b) administering the one or more senotherapeutic agents to the mammal if the mammal has the reduced level; and (c) administering, to the mammal, (i) a higher amount of the one or more senotherapeutic agents of the previous administration or (ii) one or more senotherapeutic agents that are different from the one or more senotherapeutic agents of the previous administration, if the mammal does not have the reduced level. The mammal can be a human. The determining step can include comparing the level of PAPP-A polypeptides within a first sample obtained from the mammal prior to the previous administration to the level of PAPP-A polypeptides within a second sample obtained from the mammal after the previous administration. The first and second samples can be blood samples. The first and second samples can be urine samples. The first and second samples can be saliva samples. The one or more senotherapeutic agents can be dasatinib, quercetin, navitoclax, A1331852, Al 155463, fisetin, luteolin, geldanamycin, tanespimycin, alvespimycin, piperlongumine, panobinostat, F0X04- related peptides, nutlin-3 a, ruxolitinib, metformin, or rapamycin. The method can include determining that the mammal has the reduced level. The administering the one or more senotherapeutic agents to the mammal of step (b) can include administering the one or more senotherapeutic agents to the mammal in the same manner as the previous administration. The method can include determining that the mammal does not have the reduced level. The administering of step (c) can include administering the higher amount of the one or more senotherapeutic agents of the previous administration to the mammal. The administering of step (c) can include administering the one or more senotherapeutic agents that are different from the one or more senotherapeutic agents of the previous administration. In another aspect, this document features methods for continuing a previous antisenescence treatment. The methods can include, consist of, or consist essentially of, administering an anti-senescence treatment that can be the same as a previous antisenescence treatment to a mammal identified as having a reduced level of PAPP- A polypeptides following the previous administration as compared to the level of PAPP -A polypeptides within the mammal prior to the previous administration. The mammal can be a human. The mammal can be identified as having the reduced level by comparing the level of PAPP-A polypeptides within a first sample obtained from the mammal prior to the previous anti- senescence treatment to the level of PAPP-A polypeptides within a second sample obtained from the mammal after the previous anti-senescence treatment. The first and second samples can be blood samples. The first and second samples can be urine samples. The first and second samples can be saliva samples. The one or more senotherapeutic agents can be dasatinib, quercetin, navitoclax, A1331852, Al 155463, fisetin, luteolin, geldanamycin, tanespimycin, alvespimycin, piperlongumine, panobinostat, FOXO4-related peptides, nutlin- 3a, ruxolitinib, metformin, or rapamycin.

In another aspect, this document features methods for changing an ineffective use of a previous anti-senescence treatment. The methods can include, consist of, or consist essentially of, administering an anti-senescence treatment that can be different from a previous anti-senescence treatment to a mammal identified as not having a reduced level of PAPP-A polypeptides following the previous administration as compared to the level of PAPP-A polypeptides within the mammal prior to the previous administration. The mammal can be a human. The mammal can be identified as not having the reduced level by comparing the level of PAPP-A polypeptides within a first sample obtained from the mammal prior to the previous anti-senescence treatment to the level of PAPP-A polypeptides within a second sample obtained from the mammal after the previous anti-senescence treatment. The first and second samples can be blood samples. The first and second samples can be urine samples. The first and second samples can be saliva samples. The one or more senotherapeutic agents can be dasatinib, quercetin, navitoclax, A1331852, Al 155463, fisetin, luteolin, geldanamycin, tanespimycin, alvespimycin, piperlongumine, panobinostat, FOX04-related peptides, nutlin-3a, ruxolitinib, metformin, or rapamycin. In another aspect, this document features methods for reducing the level of a PAPP -A polypeptide within a mammal. The methods can include, consist of, or consist essentially of, administering one or more senotherapeutic agents to a mammal. The mammal can be a human. The mammal can be identified as being in need of a reduced level of PAPP-A polypeptides. The one or more senotherapeutic agents can be dasatinib, quercetin, navitoclax, A1331852, Al 155463, fisetin, luteolin, geldanamycin, tanespimycin, alvespimycin, piperlongumine, panobinostat, FOXO4-related peptides, nutlin-3a, ruxolitinib, metformin, or rapamycin. The method can include determining, after the administering step, that the level of PAPP-A polypeptides was reduced within the mammal following the administering step. The determining step can include comparing the level of PAPP-A polypeptides within a first sample obtained from the mammal prior to the administering step to the level of PAPP-A polypeptides within a second sample obtained from the mammal after the administering step. The first and second samples can be blood samples. The first and second samples can be urine samples. The first and second samples can be saliva samples.

In another aspect, this document features senotherapeutic agents for use in the preparation of a medicament to reduce the level of PAPP-A polypeptides within a mammal. The mammal can be a human

In another aspect, this document features senotherapeutic agents for use in reducing the level of PAPP-A polypeptides within a mammal. The mammal can be a human.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Although methods and materials similar or equivalent to those described herein can be used to practice the invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.

The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and drawings, and from the claims. DESCRIPTION OF THE DRAWINGS

Figures 1 A - IB show that PAPP-A polypeptide expression and IGF-1 polypeptide expression were increased in senescent human adipose-derived progenitors (preadipocytes; Figure 1A) and in senescent human umbilical vein endothelial cells (HUVECs; Figure IB) relative to non-senescent cells. The amount of PAPP-A and IGF-1 RNA was assessed by relative RT-PCR in non-senescent cells and in cells made senescent by ionizing radiation.

Figures 2A - 2B show that senolytic drugs reduced secretion of PAPP-A polypeptides from human adipose tissue. Adipose tissue biopsies from human subjects (N=5) were placed in organ culture dishes and were treated with vehicle or senolytic dmgs (dasatinib+quercetin (DQ) or Fisetin (FIS)). Figure 2A is a graph of a level of PAPP-A polypeptides measured by ELISA and normalized to total protein in CM. Data are expressed as a ratio to vehicle-treated tissue. Statistical analysis used a one-way ANOVA, P<0.009 and Tukey multiple comparisons *P<0.048, **P<0.008. Tissue was digested, and PAPP-A mRNA was measured by relative RT-PCR. A TATA box binding protein (TBP) was used as a control. PAPP-A polypeptide expression was standardized to the PAPP-A level in vehicle-treated tissue. *P<0.0018.

Figure 3 shows paracrine effects of senolytic drugs on regulators of PAPP-A polypeptide expression in human adipose tissue. Adipose tissue biopsies from human subjects (N=5) were placed in organ culture dishes and were treated with vehicle or senolytic drugs (DQ or FIS). mRNA expression of pl 6, TL-la, TL-i p, and TL6 polypeptides was measured by relative RT-PCR. Fold change of mRNA expression in senolytic-treated cells was standardized to PAPP-A expression in the vehicle-treated cells and are shown in the heat map.

DETAILED DESCRIPTION

This document provides methods and materials for assessing anti-senescence treatments as well as methods and materials for effectively using anti-senescence treatments within mammals. For example, this document provides methods and materials for determining whether or not a particular anti-senescence treatment administered to a mammal (e g., a human) is effective (e g., is effective to reduce the number of senescent cells within that mammal). In some cases, determining that a mammal (e.g., a human) has a reduced level of PAPP- A polypeptides following administration of an anti-senescence treatment as compared to the level of PAPP- A polypeptides prior to being administered the antisenescence treatment can indicate that that anti-senescence treatment is effective in that mammal and can be continued as is (or with one or more slight alterations) in that mammal. In some cases, determining that a mammal (e.g., a human) does not have a reduced level of PAPP-A polypeptides following administration of an anti-senescence treatment as compared to the level of PAPP- A polypeptides prior to being administered the anti-senescence treatment can indicate that that anti-senescence treatment is not effective in that mammal and can be discontinued, altered in amount and/or frequency, or changed to a different type of anti-senescence treatment in that mammal.

Any appropriate mammal can be administered an anti-senescence treatment and have its effectiveness assessed as described herein. Any appropriate mammal that was administered or instructed to self-administer an anti-senescence treatment can be assessed as described herein to determine if the anti-senescence treatment is effective in that mammal. Any appropriate mammal can be treated with one or more anti-senescence treatments that can be assessed as described herein to arrive at a particular anti-senescence treatment that is determined to be effective in that mammal. Examples of mammals that can be assessed and/or treated as described herein include, without limitation, humans, non-human primates such as monkeys, dogs, cats, horses, cows, pigs, sheep, mice, and rats. When assessing and/or treating a mammal (e g., a human) as described herein, the mammal can be any appropriate age. For example, when assessing and/or treating a human as described herein, the human can be an adult that is at least 18 years of age (e.g., about 19, about 25, about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90 years of age).

A mammal (e.g., a human) having been administered an anti-senescence treatment (e.g., having been administered one or more senotherapeutic agents) can be assessed to determine and/or classify the efficacy of the anti-senescence treatment. For example, a mammal (e.g., a human) having been administered an anti-senescence treatment (e.g., having been administered one or more senotherapeutic agents) can be assessed to determine whether or not the anti-senescence treatment is effective within that mammal by comparing the level of PAPP-A polypeptides in a sample (e.g., a blood sample, a urine sample, or a saliva sample) obtained from the mammal following the anti- senescence treatment to the level of PAPP-A polypeptides in a comparable sample obtained from the mammal prior to the anti-senescence treatment. As described herein, senescent cells within a mammal (e.g., a human) can secrete PAPP-A polypeptides that can be detected in that mammal’s bodily fluids (e.g., in that mammal’s blood, urine, and/or saliva) and/or tissues (e.g., adipose tissues). Accordingly, levels of PAPP-A polypeptides can be used to determine whether the number of senescent cells in a mammal (e.g., a human) are increasing, decreasing, or staying the same or essentially the same, and can therefore be used to determine and/or classify whether or not an anti-senescence treatment is effective (e.g., whether or not an anti-senescence treatment reduced the number of senescent cells within the mammal).

If an anti- senescence treatment is determined to be effective, the same antisenescence treatment can be administered to the mammal (e.g., human). In some cases, if an anti-senescence treatment is determined to be effective, an anti-senescence treatment that is a slight modification of the original anti-senescence treatment can be administered to the mammal (e.g., human). For example, when an anti-senescence treatment is determined to be effective within a mammal, that mammal can be administered or instructed to self-administer a slightly higher dose (e.g., l.lx or 1.2x that of the original dose) or a slightly lower dose (e g., 0.9x or 0.8x that of the original dose) of the same agents of the effective antisenescence treatment.

If an anti-senescent treatment is determined and/or classified to not be effective, a revised anti-senescence treatment can be administered. For example, when an antisenescence treatment is determined not to be effective within a mammal, that mammal can be administered or instructed to self-administer a revised anti-senescence treatment that includes (a) an anti-senescence treatment that is different from the previously administered antisenescence treatment(s), (b) an anti- senescence treatment that is of a higher amount of the previously administered one or more senotherapeutic agents of the anti-senescence treatment, and/or (c) an anti-senescence treatment that includes one or more senotherapeutic agents that are different from the one or more senotherapeutic agents of previously administered antisenescence treatment. Any of the methods described herein used to determine if an anti- senescence treatment is effective also can be used to determine if a revised anti- senescence treatment is effective.

In some cases, an anti-senescence treatment that is different from the previously administered anti-senescence treatment can include administering one or more different senotherapeutic agents. Different anti-senescence treatments also can include administering the previously administered one or more senotherapeutic agents and an additional one or more senotherapeutic agent that was not previously administered. In some cases, an antisenescence treatment that is of a higher amount of the previously administered one or more senotherapeutic agents of the anti-senescence treatment can include an increased dosage, a more frequent dosage regime, or both. Increased dosage can include increasing the dosage by, for example, 1.3x, 1.4x, 1.5x, 1.75x, 2x, or more compared to the previous dosage of the administered senotherapeutic agent(s). More frequent dosage regimes can include administering the anti-senescence treatment more frequently (e.g., once a week, once every two weeks, once a month, every 2 months, every 3 months, every 4 months, every 5 months, or every six months) as compared to the previous regime of the administered senotherapeutic agent(s).

In some cases, a sample (e.g., a blood sample, a urine sample, or a saliva sample) can be obtained from a mammal (e.g., a human) having been administered an anti-senescence treatment (e.g., having been administered one or more senotherapeutic agents) to determine whether or not the anti-senescence treatment is effective within that mammal. In some cases, the level of PAPP- A polypeptides in the sample obtained from a mammal after the mammal was administered an anti-senescence treatment can be compared to the level of PAPP -A polypeptides within a comparable sample obtained from the mammal prior to the mammal being administered the anti-senescence treatment. In some cases, the level of PAPP-A polypeptides in the sample obtained from a mammal after the mammal was administered an anti-senescence treatment can be compared to a cutoff level of PAPP- A polypeptides that can be used to identify reduced levels of PAPP-A polypeptides that are indicative of an effective anti-senescence treatment in that mammal. For humans, a cutoff level of PAPP-A polypeptides that can be used to identify reduced levels of PAPP- A polypeptides that are indicative of an effective anti-senescence treatment in that mammal can be about 30 nanogram (ng) per milliliter (mL) (e.g., from about 19 ng per mL to about 40 ng per mL, from about 19 ng per mL to about 35 ng per mL, from about 19 ng per mL to about 30 ng per mL, from about 19 ng per mL to about 25 ng per mL, from about 25 ng per mL to about 40 ng per mL, from about 30 ng per mL to about 40 ng per mL, from about 35 ng per mL to about 40 ng per mL, from about 20 ng per mL to about 35 ng per mL, from about 23 ng per mL to about 32 ng per mL, from about 20 ng per mL to about 25 ng per mL, from about 25 ng per mL to about 30 ng per mL, or from about 30 ng per mL to about 35 ng per mL) of whole blood. For example, a cutoff level of PAPP-A polypeptides that can be used to identify reduced levels of PAPP- A polypeptides that are indicative of an effective anti- senescence treatment in a male human can be about 23 ng per mL (e.g., can be from about 19 ng per mL to about 24 ng per mL, from about 19 ng per mL to about 22 ng per mL, from about 19 ng per mL to about 20 ng per mL, from about 20 ng per mL to about 22 ng per mL, from about 19 ng per mL to about 21 ng per mL, or from about 20 ng per mL to about 22 ng per mL) of whole blood. For example, a cutoff level of PAPP-A polypeptides that can be used to identify reduced levels of PAPP- A polypeptides that are indicative of an effective anti- senescence treatment in a female human can be about 34 ng per mL (e.g., can be from about 25 ng per mL to about 37 ng per mL, from about 25 ng per mL to about 35 ng per mL, from about 25 ng per mL to about 30 ng per mL, from about 25 ng per mL to about 28 ng per mL, from about 28 ng per mL to about 37 ng per mL, from about 30 ng per mL to about 37 ng per mL, from about 33 ng per mL to about 37 ng per mL, from about 28 ng per mL to about 35 ng per mL, from about 30 ng per mL to about 33 ng per mL, from about 27 ng per mL to about 30 ng per mL, or from about 32 ng per mL to about 34 ng per mL) of whole blood. In some cases, a cutoff level of PAPP-A polypeptides that can be used to identify reduced levels of PAPP-A polypeptides that are indicative of an effective anti-senescence treatment in a mammal can be as described elsewhere (e.g., Coskun et al., (2013) Journal of Cardiology. 61: 128-131, at, for example, the abstract).

A sample (e.g., a blood sample, a urine sample, or a saliva sample) can be obtained from a mammal (e.g., a human) before and/or after the mammal has been administered an anti-senescence treatment (e.g., one or more senotherapeutic agents). In some cases, a sample (e g., a blood sample, a urine sample, or a saliva sample) can be obtained from a mammal (e g., a human) at any appropriate time before the mammal has been administered an antisenescence treatment and assessed to determine the level of PAPP -A polypeptides within the mammal prior to being administered the anti-senescence treatment. For example, a sample (e.g., a blood sample, a urine sample, or a saliva sample) can be obtained from a mammal (e.g., a human) within five minutes, 30 minutes, one hour, six hours, 12 hours, one day, two days, three days, five days, one week, two weeks, three weeks, four weeks, one month, or two months before the mammal has been administered an anti-senescence treatment and assessed to determine the level of PAPP- A polypeptides within the mammal prior to being administered the anti-senescence treatment. Such a determined level of PAPP-A polypeptides within the mammal prior to being administered the anti- senescence treatment can be used to determine if the level of PAPP- A polypeptides within the mammal determined after the mammal has been administered an anti-senescence treatment is reduced, is elevated, or remains the same following the anti-senescence treatment. A reduction in the level of PAPP- A polypeptides within the mammal after the mammal has been administered an antisenescence treatment can indicate that an anti-senescence treatment is effective in that mammal.

In some cases, a sample (e.g., a blood sample, a urine sample, or a saliva sample) can be obtained from mammal (e.g., a human) during the time that the mammal is administered an anti-senescence treatment and assessed to determine the level of PAPP-A polypeptides within the mammal at that time. For example, a sample (e g., a blood sample, a urine sample, or a saliva sample) can be obtained from a mammal (e.g., a human) at the same time (e.g., at the start of an anti-senescence treatment) that the mammal is being administered an antisenescence treatment and assessed to determine the level of PAPP -A polypeptides within the mammal at that time. Such a determined level of PAPP- A polypeptides within the mammal at the time of administration of the anti-senescence treatment can be used to determine if the level of PAPP- A polypeptides within the mammal determined after the mammal has been exposed to the administered anti-senescence treatment for a period of time is reduced, is elevated, or remains the same following the anti-senescence treatment. A reduction in the level of PAPP- A polypeptides within the mammal after the mammal has been exposed to the administered anti-senescence treatment for a period of time can indicate that that an antisenescence treatment is effective in that mammal.

In some cases, to assess the effectiveness of an anti-senescence treatment within a mammal, a sample (e.g., a blood sample, a urine sample, or a saliva sample) can be obtained from the mammal (e.g., a human) at any appropriate time after the mammal has been administered an anti-senescence treatment and assessed to determine the level of PAPP-A polypeptides within the mammal after being administered the anti-senescence treatment. For example, a sample (e.g., a blood sample, a urine sample, or a saliva sample) can be obtained from a mammal (e.g., a human) between about 2 weeks and about 6 months (e g., between about 2 weeks and about 5 months, between about 2 weeks and about 4 months, between about 2 weeks and about 3 months, between about 2 weeks and about 2 months, between about 2 weeks and about 1 month, between about 2 weeks and about 3 weeks, between about 3 weeks and about 6 months, between about 1 month and about 6 months, between about 2 months and about 6 months, between about 3 months and about 6 months, between about 4 months and about 6 months, between about 5 months and about 6 months, between about 3 weeks and about 5 months, between about 1 month and about 4 months, or between about 2 months and about 3 months) after an initial administration of an anti-senescence treatment and assessed to determine the level of PAPP-A polypeptides within the mammal after being administered the anti-senescence treatment. Such a determined level of PAPP-A polypeptides within the mammal after being administered the anti-senescence treatment can be used to determine if the level of PAPP- A polypeptides within the mammal determined after the mammal has been administered an anti-senescence treatment is reduced, is elevated, or remains the same following the anti-senescence treatment. A reduction in the level of PAPP- A polypeptides within the mammal after the mammal has been administered an antisenescence treatment can indicate that that an anti- senescence treatment is effective in that mammal.

In some cases, the level of PAPP-A polypeptides within a mammal (e.g., within a sample obtained from a mammal) can be detected at different time points over a course of an anti-senescence treatment (and/or a revised anti- senescence treatment) to determine the efficacy of the anti-senescence treatment (and/or revised anti-senescence treatment). For example, two or more (e.g., two, three, four, five, six, or more) samples (e.g., blood samples, urine samples, or saliva samples) can be obtained from a mammal at different time points over the course of an anti-senescence treatment (and/or a revised anti-senescence treatment), and the level of PAPP-A polypeptides in the samples can be used to determine the efficacy of the anti-senescence treatment (and/or a revised anti-senescence treatment).

In some cases, a first sample (e.g., a first blood sample, a first urine sample, or a first saliva sample) can be obtained from a mammal (e.g., a human) prior to the mammal being administered an anti-senescence treatment (e.g., prior to being administered one or more senotherapeutic agents), and a second sample (e.g., a second blood sample, a second urine sample, or a second saliva sample), and optionally subsequent samples, can be obtained from the mammal after the mammal has been administered the anti-senescence treatment. When the level of PAPP- A polypeptides in the second sample is less than (e.g., 1, 2, 3, 4, 5, 10, 15, 20, 25, 50, or more percent less than) the level of PAPP-A polypeptides in the first sample, then the anti-senescence treatment (e.g., one or more senotherapeutic agents) can be determined and/or classified as being an effective anti-senescence treatment for that mammal. When the level of PAPP -A polypeptides in the second sample is greater than (e.g., 1, 2, 3, 4, 5, 10, 15, 20, 25, 50, or more percent greater than) or equal to the level of the PAPP-A polypeptides in the first sample, then the anti-senescence treatment (e g., one or more senotherapeutic agents) can be determined and/or classified as not being an effective treatment for that mammal.

Any appropriate sample from a mammal (e.g., a human) can be assessed for the presence, absence, or level of PAPP-A polypeptides. In some cases, a sample can be a biological sample. For example, a sample can contain one or more biological molecules (e.g., nucleic acids such as DNA and RNA, polypeptides, carbohydrates, lipids, hormones, and/or metabolites). Examples of samples that can be assessed to determine the level of PAPP-A polypeptides as described herein include, without limitation, fluid samples (e.g., urine samples, saliva samples, and blood samples such as whole blood samples, serum samples, and plasma samples), and tissue samples (e.g., adipose tissue, skin, muscle, heart, vessels, kidney, blood cells, extracellular vesicle tissue, and liver tissue). A biological sample can be a fresh sample or a fixed sample (e.g., a formaldehyde-fixed sample or a formalin-fixed sample). In some cases, a biological sample can be a processed sample (e.g., to isolate or extract one or more biological molecules). For example, a blood sample can be obtained from a mammal (e.g., a human) and processed to obtain a cellular sample or a polypeptide sample, which can be assessed for the presence, absence, or level of PAPP- A polypeptides as described herein.

When assessing the efficacy of an anti-senescence treatment within a mammal (e.g., a human) as described herein (e.g., by assessing the level of PAPP-A polypeptides in a sample obtained from a mammal having been administered an anti-senescence treatment), the level of any appropriate PAPP-A polypeptide can be detected. Examples of PAPP- A polypeptides that can be used as described herein include, without limitation, a human PAPP-A polypeptide having the amino acid sequence set forth in National Center for Biotechnology Information (NCBI) GenBank® or GenPept® Accession No. NP_002572.2, a monkey PAPP- A polypeptide having the amino acid sequence set forth in NCBI GenBank® or GenPept® Accession No. XP_014972456.2, a horse PAPP-A polypeptide having the amino acid sequence set forth in NCBI GenBank® or GenPept® Accession No. XP_023484521, a dog PAPP-A polypeptide having the amino acid sequence set forth in NCBI GenBank® or GenPept® Accession No. XP_038347014.1, a cat PAPP-A polypeptide having the amino acid sequence set forth in NCBI GenBank® or GenPept® Accession No. XP_003993206.1, a mouse PAPP-A polypeptide having the amino acid sequence set forth in NCBI GenBank® or GenPept® Accession No. NP_067337.1 , or a rat PAPP-A polypeptide having the amino acid sequence set forth in NCBI GenBank® or GenPept® Accession No. XP_006238331.

Any appropriate method can be used to detect the presence, absence, or level of PAPP-A polypeptides within a sample (e.g., a sample obtained from a mammal such as a human) to determine the efficacy of an anti-senescence treatment. In some cases, the presence, absence, or level of PAPP-A polypeptides within a sample can be determined by detecting the presence, absence, or level of PAPP- A polypeptides in the sample. For example, immunoassays (e.g., immunohistochemistry (IHC) techniques and western blotting techniques), mass spectrometry techniques (e.g., proteomics-based mass spectrometry assays or targeted quantification-based mass spectrometry assays), cytometry by time of flight, (CyTOF), RT-PCR, and ELISA can be used to determine the presence, absence, or level of PAPP- A polypeptides in a sample. When an immunoassay is used to determine the presence, absence, or level of PAPP- A polypeptides in a sample, the immunoassay can use any appropriate antibody. Examples of antibodies that can be used in an immunoassay to determine the presence, absence, or level of PAPP- A polypeptides in a sample include, without limitation, PAPP-A/Pappalysin- 1 monoclonal antibodies (e.g., NeoBiotechnologies, PAPPA, 2715, Cat. No. 5069-MSM1-P1), human pappalysin-l/PAPP- A antibodies (e.g., R&D Systems, Cat. No. AF2487), and PAPP-A(pico) ELISA kit antibodies (e.g., ANSH LABs, Cat. No. AL-101).

In some case, the level of mRNA encoding a PAPP-A polypeptide can be measured to indicate the level of PAPP- A polypeptides within a mammal. For example, a sample containing cells can be obtained from a mammal, and the level of mRNA encoding PAPP-A polypeptides can be measured to determine if the level of PAPP- A polypeptides within the mammal was reduced following administration of an anti-senescence treatment. Any appropriate method can be used to measure the level of mRNA encoding a PAPP-A polypeptide within a sample. For example, polymerase chain reaction (PCR)-based techniques such as quantitative reverse transcription (RT)-PCR (qRT-PCR) techniques, single cell RNA sequencing, and bulk RNA sequencing can be used to determine the presence, absence, or level of mRNA encoding a PAPP-A polypeptide in the sample. In some cases, the presence, absence, or level of mRNA encoding a PAPP-A polypeptide within a sample can be determined by qRT-PCR. Tn some cases, the presence, absence, or level of mRNA encoding a PAPP-A polypeptide within a sample can be determined as described in Example 2 and/or Example 3.

Any appropriate anti-senescence treatment (and/or revised anti-senescence treatment) can be administered to a mammal (e.g., a human) and be assessed for effectiveness as described herein (e.g., by assessing the level of PAPP-A polypeptides after the mammal has be administered the anti-senescence treatment being assessed). An anti-senescence treatment is a treatment designed to inhibit or reduce cellular senescence within a mammal. For example, an anti-senescence treatment that can be administered to a mammal (e.g., a human) and/or assessed as described herein can be a treatment that includes administering one or more senotherapeutic agents to the mammal (e.g., the human). Examples of senotherapeutic agents that can be used as at least a part of an anti- senescence treatment described herein include, without limitation, geroprotector agents such as melatonin, carnosine, metformin, rapamycin, nicotinamide mononucleotide (NMN) and delta sleep-inducing peptide, SASP (senescence-associated secretory phenotype) inhibitors such as glucocorticoids, statins (e.g., simvastatin), JAK1/2 inhibitors (e.g., ruxolitinib), NF-KB inhibitors, p38 inhibitors, and IL- la inhibitors, senolytic agents such as FOXO4-related peptides, BCL-2 inhibitors, Src inhibitors, USP7 inhibitors, quercetin plus dasatinib, fisetin, navitoclax, piperlongumine, azithromycin, roxithromycin, senescence-specific killing compound 1 (SSK1), GLS1 inhibitors, 25-hydroxycholesterol, Procyanidin Cl, and EF-24, and senomorphic agents such as apigenin, rapamycin, and everolimus. Additional examples of senotherapeutic agents that can be administered and/or assessed as described herein include, without limitation, A1331852, A1155463, luteolin, geldanamycin, tanespimycin, alvespimycin, panobinostat, nutlin-3a, and ruxolitinib. In some cases, a senotherapeutic agent that can be used as described herein can be a senotherapeutic agent as described elsewhere (see, e.g., Kirkland et al., J. Intern. Med., 288(5):518-536 (2020) at, for example, Table 1).

This document also provides methods and materials for treating a mammal (e.g., a human) in need of inhibited or reduced cellular senescence. For example, a mammal (e.g., a human) can be treated as described herein by administering any appropriate anti-senescence treatment (and/or revised anti-senescence treatment) to the mammal and by confirming the effectiveness of that anti-senescence treatment (and/or revised anti-senescence treatment) within that mammal as described herein. In such cases, the anti-senescence treatment (and/or revised anti-senescence treatment) determined to be effective within that mammal can be continued as is (or with one or more slight alterations).

In some cases, a method for treating a mammal (e.g., a human) provided herein can include administering a previously administered anti-senescence treatment (or slight modification thereof) to a mammal that received that previous anti-senescence treatment and that previous anti-senescence treatment was determined to be effective within that mammal as described herein. In some cases, that previously effective anti-senescence treatment can be continued as is. In some cases, that previously effective anti-senescence treatment can be continued with one or more slight alterations. In some cases, a method for treating a mammal (e.g., a human) provided herein can include administering a first anti-senescence treatment to the mammal, determining that that first anti-senescence treatment is not effective within that mammal as described herein, and administering a second (e.g., a revised anti-senescence treatment) to the mammal (and optionally confirming the effectiveness of that revised anti-senescence treatment within that mammal) as described herein.

In some cases, a method for treating a mammal (e.g., a human) provided herein can include administering an anti-senescence treatment (e.g., a revised anti-senescence treatment) to a mammal that received a previous anti-senescence treatment that was determined to be ineffective as described herein. In such cases, the administered an anti-senescence treatment (e g., a revised anti-senescence treatment) can be different from that of the previous antisenescence treatment that was determined to be ineffective. In some cases, the effectiveness of that administer anti-senescence treatment can be assessed as described herein.

In some cases, a method of treating a mammal (e.g., a human) to inhibit or reduce cellular senescence can include a step of determining whether or not the mammal has a reduced level of PAPP- A polypeptides following a previous administration of an antisenescence treatment to the mammal as compared to the level of PAPP -A polypeptides within the mammal prior to the previous administration. In some cases, the method can include administering the same anti-senescence treatment (or a slightly modified version thereof) to the mammal (e.g., human) if the mammal has a reduced level of a PAPP-A polypeptide as compared to the level of PAPP-A polypeptides within the mammal prior to the previous administration of an anti-senescence treatment. In some cases, the method can include administering a revised anti-senescence treatment to the mammal (e.g., human) if the mammal does not have a reduced level of a PAPP-A polypeptide as compared to the level of PAPP-A polypeptides within the mammal prior to the previous administration of an antisenescence treatment. In some cases, the revised anti-senescence treatment can be a higher dosage amount (e.g., about 1.3x to about 4x higher than the previous administration, about 1 ,4x to about 4x, about 1 ,5x to about 4x, about 1.75x to about 4x, about 2x to about 4x, about 2.5x to about 4x, about 3x to about 4x, about 1 ,3x to about 3x, about 1 ,3x to about 2.5x, about 1.3x to about 2x, about 1.3x to about 1.75x, about 1.3x to about 1.5x, about 1.3x to about 1.4x, about 1.3x, about 1 4x, about 1.5x, about 1.75x, or about 2x, about 2.5x, about 3x, about 4x) of the same senotherapeutic agent(s) of the previous administration of an antisenescence treatment as compared to the amounts of the senotherapeutic agent(s) of the previous administration of an anti- senescence treatment. In some cases, the revised antisenescence treatment can be a more frequent administration (e.g., once a week, once every two weeks, once a month, every 2 months, every 3 months, every 4 months, every 5 months, or every six months) of the same senotherapeutic agent(s) of the previous administration of an anti-senescence treatment as compared to the amounts of the senotherapeutic agent(s) of the previous administration of an anti-senescence treatment. In some cases, the revised antisenescence treatment can be designed to include one or more senotherapeutic agents that are different from the senotherapeutic agent(s) of the previous administration of an antisenescence treatment.

Any appropriate anti-senescence treatment, revised anti- senescence treatment, or senotherapeutic agent described herein can be administered to a mammal (e.g., a human) to inhibit or reduce cellular senescence within a mammal and/or to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease.

In some cases, an anti-senescence treatment (e.g., one or more senotherapeutic agents) can be formulated into a composition (e.g., a pharmaceutically acceptable composition) for administration to a mammal having a disease or disorder characterized by excess cellular senescence (e.g., an age-related disease). For example, one or more senotherapeutic agents can be formulated together with one or more pharmaceutically acceptable carriers (additives), excipients, and/or diluents. In some cases, a pharmaceutically acceptable carrier, excipient, or diluent can be a naturally occurring pharmaceutically acceptable carrier, excipient, or diluent. In some cases, a pharmaceutically acceptable carrier, excipient, or diluent can be a non-naturally occurring (e.g., an artificial or synthetic) pharmaceutically acceptable carrier, excipient, or diluent. Examples of pharmaceutically acceptable carriers, excipients, and diluents that can be used in a composition described herein include, without limitation, serum proteins (e.g., human serum albumin), water, polyethylene glycol, salts and electrolytes (e.g., phosphate salts, saline, protamine sulfate, and DMSO).

An anti-senescence treatment (e.g., one or more senotherapeutic agents) can be administered to a mammal (e.g., a human) by any appropriate route (e.g., oral, intranasal, inhalation, transdermal, and parenteral). An anti-senescence treatment can be administered to a mammal locally or systemically. For example, an anti- senescence treatment can be administered locally by topical administration (e.g., as an ointment) to a mammal (e.g., a human). For example, an anti-senescence treatment can be administered locally by injection to a mammal (e.g., a human). For example, an anti-senescence treatment can be administered systemically by oral administration to a mammal (e.g., a human).

An anti-senescence treatment (e.g., one or more senotherapeutic agents) can be administered to a mammal (e.g., a human) in any appropriate amount (e.g., any appropriate dose). An effective amount of an anti-senescence treatment can be any amount that reduces the level of PAPP- A polypeptides with the mammal being treated without producing significant toxicity to the mammal. In cases where an anti-senescence treatment includes dasatinib, an effective amount (e.g., an effective daily amount) of dasatinib can be from about 1.5 milligrams per kilogram body weight (mg/kg) to about 3 mg/kg(e.g. from about 1.5 mg/kg to about 2.8 mg/kg, from about 1.5 mg/kg to about 2.5 mg/kg, from about 1.5 mg/kg to about 2.2 mg/kg, from about 1.5 mg/kg to about 2 mg/kg, from about 1.8 mg/kg to about 3 mg/kg, from about 2 mg/kg to about 3 mg/kg, from about 2.3 mg/kg to about 3 mg/kg, from about 2.5 mg/kg to about 3 mg/kg, from about 1.8 mg/kg to about 2.5 mg/kg, from about 2 mg/kg to about 2.3 mg/kg, from about 1.8 mg/kg to about 2 mg/kg, from about 2 mg/kg to about 2.5 mg/kg, or from about 2.5 mg/kg to about 2.8 mg/kg). In cases where an antisenescence treatment includes quercetin, an effective amount (e.g., an effective daily amount) of quercetin can be from about 15 mg of quercetin per kg of body weight (mg/kg) to about 30 mg/kg (e.g., from about 15 mg/kg to about 25 mg/kg, from about 15 mg/kg to about 20 mg/kg, from about 15 mg/kg to about 18 mg/kg, from about 20 mg/kg to about 30 mg/kg, from about 25 mg/kg to about 30 mg/kg, from about 28 mg/kg to about 30 mg/kg, from about 18 mg/kg to about 25 mg/kg, from about 20 mg/kg to about 22 mg/kg, from about 18 mg/kg to about 20 mg/kg, from about 20 mg/kg to about 25 mg/kg, or from about 25 mg/kg to about 28 mg/kg). In cases where an anti-senescence treatment includes fisetin, an effective amount (e.g., an effective daily amount) of fisetin can be from about 20 mg of fisetin per kg of body weight (mg/kg) to about 100 mg/kg (e.g from about 20 mg/kg to about 80 mg/kg, from about 20 mg/kg to about 60 mg/kg, from about 20 mg/kg to about 50 mg/kg, from about 20 mg/kg to about 40 mg/kg, from about 20 mg/kg to about 30 mg/kg, from about 40 mg/kg to about 100 mg/kg, from about 60 mg/kg to about 100 mg/kg, from about 80 mg/kg to about 100 mg/kg, from about 30 mg/kg to about 90 mg/kg, from about 40 mg/kg to about 80 mg/kg, from about 50 mg/kg to about 70 mg/kg, from about 30 mg/kg to about 50 mg/kg, from about 40 mg/kg to about 60 mg/kg, from about 60 mg/kg to about 80 mg/kg, or from about 70 mg/kg to about 90 mg/kg). The effective amount can remain constant or can be adjusted as a sliding scale or variable dose depending on the mammal’s response to treatment. Various factors can influence the actual effective amount used for a particular application. For example, the frequency of administration, duration of treatment, use of multiple treatment agents, route of administration, and/or severity of the disease or disorder characterized by excess cellular senescence (e.g., an age-related disease) in the mammal being treated may require an increase or decrease in the actual effective amount administered.

An anti-senescence treatment (e.g., one or more senotherapeutic agents) can be administered to a mammal (e.g., a human) at any appropriate frequency. The frequency of administration can be any frequency that reduces the level of PAPP- A polypeptides with the mammal being treated without producing significant toxicity to the mammal. For example, the frequency of administration can be from about twice a day to about one every other day, from about once a day to about once a week, from about once a day to about once a month, from about once a week to about once a month, or from about twice a month to about once a month. In some cases, the frequency of administration can be daily for one or more (e.g., one, two, three, four, or more) consecutive days. The frequency of administration can remain constant or can be variable during the duration of treatment. As with the effective amount, various factors can influence the actual frequency of administration used for a particular application. For example, the effective amount, duration of treatment, use of multiple treatment agents, and/or route of administration may require an increase or decrease in administration frequency. An anti-senescence treatment (e.g., one or more senotherapeutic agents) can be administered to a mammal (e.g., a human) for any appropriate duration. An effective duration for administering or using an anti-senescence treatment can be any duration that reduces the level of PAPP- A polypeptides with the mammal being treated without producing significant toxicity to the mammal. For example, the effective duration can vary from several weeks to several months, from several months to several years, or from several years to a lifetime. Multiple factors can influence the actual effective duration used for a particular treatment. For example, an effective duration can vary with the frequency of administration, effective amount, use of multiple treatment agents, and/or route of administration.

In some cases, methods for treating a mammal (e g., a human) as described herein can include administering to the mammal one or more (e.g., one, two, three, four, or more) senotherapeutic agents as the sole active ingredient. For example, a composition containing one or more senotherapeutic agents can include the one or more senotherapeutic agents as the sole active ingredient in the composition for inhibiting or reducing cellular senescence.

In some cases, methods for treating a mammal (e g., a human) as described herein also can include administering to the mammal one or more (e.g., one, two, three, or more) additional agents used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease and/or performing one or more (e.g., one, two, three, or more) therapies used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease on the mammal. For example, a combination therapy used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease can include administering to the mammal (e.g., a human) one or more senotherapeutic agents as described herein and one or more (e.g., one, two, three, or more) additional agents used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease. In some cases, an additional agent that can be used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease can be an anti-fibrotic agent. Examples of additional agents that can be administered to a mammal to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease include, without limitation, metformin, losartan, rapamycin, and any combinations thereof. In cases where one or more senotherapeutic agents are used in combination with additional agents used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease, the one or more additional agents can be administered at the same time (e.g., in a single composition containing both one or more senotherapeutic agents and the one or more additional agents) or independently. For example, one or more senotherapeutic agents described herein can be administered first, and the one or more additional agents administered second, or vice versa.

In some cases, a combination therapy used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease can include administering to the mammal (e.g., a human) one or more (e.g., one, two, three, or more) senotherapeutic agents described herein and performing one or more (e.g., one, two, three, or more) additional therapies used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease include, without limitation, lifestyle interventions, diet interventions, exercise, anti-smoking interventions, and/or stress reduction. In cases where one or more senotherapeutic agents described herein are used in combination with one or more additional therapies used to treat a mammal having a disease or disorder characterized by excess cellular senescence such as an age-related disease, the one or more additional therapies can be performed at the same time or independently of the administration of one or more senotherapeutic agents described herein. For example, one or more senotherapeutic agents described herein can be administered before, during, or after the one or more additional therapies are performed.

In some cases, a course of treatment can be monitored as described herein. For example, the levels of PAPP-A polypeptides within the mammal can be monitored over the course of treatment to determine whether or not the treatment is effective and/or remains effective over time.

The invention will be further described in the following examples, which do not limit the scope of the invention described in the claims. EXAMPLES

Example 1: PAPP -A expression in senescent cells

Non-senescent human adipose-derived progenitors (preadipocytes) were isolated from subcutaneous adipose tissue from healthy kidney donors, and human umbilical vein endothelial cells (HUVECs) were made senescent by exposing the cells to 10 Gy ionizing radiation. Preadipocytes became senescent after 14 days, and HUVECs became senescent after 20 days. Senescence was assessed by staining for SA beta GAL activity. Sham irradiated cells were used as control. Cells were collected, RNA was isolated, and gene expression was measured by RT- PCR. Data were normalized to housekeeping gene TATA- binding protein expression and presented as a fold increase relative to the expression in nonsenescent cells.

Both PAPP-A polypeptide expression and IGF-1 polypeptide expression were increased in senescent pre-adipocytes and in senescent HUVECs compared to non-senescent cells (Figure 1). Graphs represent means ± SEM, T-test, asterisks:. *P < 0.01, ***P < 0.001.

Example 2: Senolytics and PAPP-A polypeptide secretion

To determine the effect of senolytics on the secretion of PAPP-A from human adipose tissue, greater omental adipose tissue was obtained from obese human subjects. Adipose tissue was cut into small pieces and washed with PBS 3 times. Adipose tissue was then cultured in medium containing 1 mM sodium pyruvate, 2 mM glutamine, MEM vitamins, MEM non-essential amino acids, and antibiotics with 20 pM Q + 1 pM D, 20 pM Fm or DMSO. After approximately 48 hours, the adipose explants were washed 3 times with PBS. Aliquots of adipose tissue were fixed for immunostaining or SA-Pgal assay. The rest of the tissue was maintained in the same medium without drugs for 24 hours to collect conditioned medium (CM). PAPP-A was measured in CM by ELISA and normalized to total protein in CM. Data are expressed as a ratio to vehicle-treated tissue. MEN±SEM, One -way ANOVA, P<0.009, Tukey multiple comparisons *P<0.048, **P<0.008 (Figure 2A). After collection of CM, adipose tissue was digested, and PAPP-A mRNA expression was measured by relative RT-PCR. The housekeeping gene TBP was used as a control. PAPPA-expression in treatment groups is expressed as a ratio to PAPPA-A level in vehicle-treated tissue. MEN±SEM, One -way ANOVA **P<0.0018 (Figure 2B).

Example 3: Senolytics and regulators of PAPP -A expression

Adipose tissue biopsies from greater omentum of obese human subjects (N=5) were placed in organ culture dishes and were treated with vehicle or senolytic drugs (DQ or fisetin). After 48 hours of treatment, tissues were washed and provided with fresh media, and CM was collected after 24 hours (as described in Example 2). RNA was collected from cultured adipose tissue biopsies, and mRNA expression of pl 6, IL- la, IL-ip, and IL6 (known regulators of PAPPA-A expression) were measured by relative RT-PCR.

The heat map represents fold change of mRNA expression of senescence marker pl6 and known regulators of PAPPA-A expression (IL- la, IL-ip, and IL6) compared to those levels in the vehicle sample (Figure 3).

Example 4: Assessing the Efficacy of an Anti-Senescence Treatment

A first blood sample is obtained from a human prior to the human being administered an anti-senescence treatment (e.g., prior to being administered one or more senotherapeutic agents), and a second blood sample is obtained from the human after the human has been administered the anti-senescence treatment.

The first and second blood samples are assessed to determine whether or not the level of PAPP- A polypeptides present in the second blood sample was less than that of the first blood sample.

When the level of PAPP-A polypeptides in the second blood sample is less than the level of PAPP-A polypeptides in the first blood sample, then the anti-senescence treatment (e g., one or more senotherapeutic agents) can be determined to be an effective treatment for that human. In this case, the human can continue with the same anti-senescence treatment.

When the level of PAPP -A polypeptides in the second blood sample is not less than the level of PAPP-A polypeptides in the first blood sample, then the anti-senescence treatment (e g., one or more senotherapeutic agents) can be determined to not be an effective treatment for that human. In this case, the human can discontinue that particular anti- senescence treatment. Optionally, that human also can initiate an increased dosing regimen of that particular anti- senescence treatment or can initiate a different anti-senescence treatment.

Example 5: Assessing the Efficacy of an Anti -Senescence Treatment

A first urine sample is obtained from a human prior to the human being administered an anti-senescence treatment (e.g., prior to being administered one or more senotherapeutic agents), and a second urine sample is obtained from the human after the human has been administered the anti-senescence treatment.

The first and second urine samples are assessed to determine whether or not the level of PAPP-A polypeptides present in the second urine sample was less than that of the first urine sample.

When the level of PAPP -A polypeptides in the second urine sample is less than the level of PAPP-A polypeptides in the first urine sample, then the anti-senescence treatment (e g., one or more senotherapeutic agents) can be determined to be an effective treatment for that human. In this case, the human can continue with the same anti-senescence treatment.

When the level of PAPP-A polypeptides in the second urine sample is not less than the level of PAPP- A polypeptides in the first urine sample, then the anti-senescence treatment (e g., one or more senotherapeutic agents) can be determined to not be an effective treatment for that human. In this case, the human can discontinue that particular anti- senescence treatment. Optionally, that human also can initiate an increased dosing regimen of that particular anti-senescence treatment or can initiate a different anti-senescence treatment.

OTHER EMBODIMENTS

It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.